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Band-in-a-Box^® 2021 for Mac Update - Build 511 (100 MB)

This patch will update Band-in-a-Box^® 2021 to build 511 from any previous 2021 build. This includes DAW Plugin 3.7.23. You need to have Band-in-a-Box^® 2021 (build 502 or higher) installed for this update to work.

If you are updating an external PG Music hard drive, make sure to choose the correct destination volume ([Change Install Location] button / Install on a specific disk. NOTE: if you get a message that it must be installed on the system drive, close other running applications and start the installer again, or if that does not resolve it, reboot the Mac.

Note: If you have a localized version (German, French, Italian), you should not install this patch, but should instead download the latest patch from here.

Summary of changes for build 511 (July 29, 2021)

• Fixed: The MIDI keyboard wizard might not work during recording.
• Fixed: "flat, dry, and center" option in the Drop Plus dialog should be available for dragging single tracks.
• Fixed: MIDI files containing sysex data would fail to import.
• Fixed: Dropping a MIDI file onto Band-in-a-Box would be ignored if the file extension was ".midi".
• Fixed: Mouse wheel scrolling did not work in the MIDI Import dialog.
• Fixed: When importing MIDI files "Merge with existing data" did not work correctly.
• Fixed: Undoing audio edits on tracks Utility#2 or higher did not work.
• Fixed: Cut/Copy/Paste hot keys did not work in Open/Save dialogs if launched from a modal dialog (eg the Render dialog).
• Fixed: When asked to overwrite song file [No] should be the default button (activated by Return key).
• Fixed: The [Cancel] button in popup dialogs should selectable with tab and arrow keys (activated by Space Bar).
• Fixed: Erase Track causes tracks to change solo state.

Summary of changes for DAW Plugin 3.7.23 (July 29, 2021)

• Fixed: VST3 did not correctly load the dedicated synth
• Fixed: "Set default style" not working
• Fixed: Moving files when saving doesn't work if style is loaded but not generated
• Fixed: DLS Music Device not loading in VST3.
• Added: Preview N Bars to chord sheet right-click menu
• Added: Message notifying that saved window size has completed
• Added: Bar highlighting for midi preview
• Fixed: Clicking anywhere on plugin while bbm4 window is open should bring window to front (plugin will now be greyed out while bb dialog is open)
• Fixed: Change disabled Preview N Bars to Preview 4 Bars by default
• Fixed: Chord sheet/track table view should not resize if horizontal size has been changed
• Fixed: Some strings not externalized in chord builder
• Fixed: PLAY/STOP buttons should be green/black to match BIAB
• Fixed: Gain value changes when toggling volume slider
• Fixed: "Move media to project directory" when saving was not always working
• Fixed: Limit for choruses and start bar were incorrect
• Fixed: Selecting secondary chord display and then going back to none doesn't resize rows well
• Fixed: Components incorrectly overlaid in confirmation dialog
• Fixed: Expand clear renders feature to exclude currently generated tracks
• Fixed: Expand auto-clear feature to work when starting new session
• Fixed: Plugin sometimes resets when closing/reopening plugin editor
• Fixed: "Auto-generate song" was not working with song demos
• Fixed: noise when starting sync playback in the plugin
• Fixed: chord sheet should get keyboard focus after any action
• Fixed: generate "region" not working
• Fixed: Save Window Size was not working

Summary of changes for build 509 (July 08, 2021)

• Fixed: Plugins not saved correctly in song files.
• Fixed: Frozen RealDrums RealCharts were not saved in song files.
• Fixed: Issues with different tempo % changes at different choruses, and starting playback in different choruses.

Summary of changes for build 507

• Fixed: The time signature does not update on the chord sheet after selecting a style.
• Fixed: Using mouse wheel over Track Selector control does not work to select None (eg in Record dialog).
• Fixed: Drag and drop Master track to MIDI quadrant, and 1 track is soloed, should be SMF type 0 instead of 1.
• Fixed: Band-in-a-Box should automatically copy Factory_stylz1.in to stylz1.bin if stylz1.bin is missing.
• Fixed: File > Revert to saved should not ask user if they want to save their current song.
• Fixed: Issues with chord sheet scrolling when near end of song.
• Fixed: Cancel should be triggered with [Esc] key in the post record audio dialog.
• Fixed: Plugins on Utility tracks do not save in song files.
• Fixed: Track Actions > Save as performance track would fail.
• Fixed: Using Copy/Move Tracks to copy audio to a non-editable track (eg Soloist) would fail.
• Fixed: Possible crash when opening the Notation window.
• Fixed: When choosing a different language the message "New Language will take effect the next time you run the program." should be displayed in the new language as well as the current language.
• Fixed: Key sig being drawn with the wrong font and position.
• Fixed: Drum notes in bottom staff that were saved to musicXML would display way too low when loaded into other programs.
• Added: Feature Browser menu item added to the Help menu.

Summary of changes for build 506 (June 29, 2021)

• Added: "Avoid connecting to low quality input devices" setting added to Audio Devices dialog.
• Fixed: Audio Unit plugins (eg the default synth) might be silent with some Audio Devices (eg bluetooth headphones).
• Fixed: Delete Beats at Bar was not undoable.
• Fixed: Delete Beats at Bar should ask for chorus/bar numbers rather than just a bar number.
• Fixed: Insert Beats at Bar should ask for chorus/bar numbers rather than just a bar number.
• Fixed: Chords and bar lines might not line up properly in the Piano Roll.
• Fixed: "Utility Tracks were automatically transposed." message shows after leaving song settings when no transpose settings were changed.

Summary of changes for build 505

• Fixed: Memory leaks when regenerating RealTracks or playing songs several times in a program session.
• Fixed: Styles that contain loops might try to load a WMA file and crash the program.
• Fixed: Quick render menu items should only render the song mix, and ignore "One file per track" setting.
• Fixed: Quick render menu items were ignoring the "Save in current song directory" setting.
• Fixed: Track context menu > Track Actions > Save track as WAV file should select stereo/mono automatically.
• Fixed: "Also generate MIDI Drums in the MIDI file" did not export the underlying MIDI style if the song has RealDrums RealCharts.
• Fixed: New Xtra Style might not display correctly in the StylePicker.
• Fixed: If scanning or loading a VST plugin causes Band-in-a-Box to crash, then that plugin should be automatically excluded the next time running the program.
• Added: Plugin selector dialog shows info on listed plugins, such as the file path and unique identifier. This is useful for troubleshooting plugin installation problems, duplicates, etc.
• Added: "Allow multiple instances of Band-in-a-Box to be running" setting added to the Preferences dialog.
• Added: [MIDI file] button added to the Preferences dialog. This replaces all of the check boxes related to MIDI exporting.
• Added: Batch Mode for exporting MIDI files is now accessible from the MIDI export dialog.
• Improved: Audio Devices dialog is resizable.

• Fixed: Partially hidden check boxes in the MIDI File Options dialog.
• Fixed: Batch Mode button should not be showing in the MIDI export dialog.
• Fixed: Drag and drop render might cause an hour glass mouse cursor to get stuck.
• Fixed: Drag and drop individual MIDI tracks should be SMF type 0 instead of 1.
• Fixed: Frozen RealTracks on Drums track are not saved in song.
• Fixed: VST plugin scans may be slower in some cases.
• Fixed: It was not possible to select audio inputs on multi-channel audio interfaces.
• Fixed: New Xtra styles do not show in the StylePicker for localized versions if the language patch is not up to date.
• Improved: File rendering time reduced.
• Improved: Enabling render setting "Individual tracks flat, dry, and center" will improve rendering time.
• Added: "Include Mix" option added to rendering options which can be disabled if rendering individual tracks.
• Added: "Save in current song directory" setting added to Render dialog.
• Added: MIDI Chord Detection window added. This can be accessed from Window > MIDI Chord Detection...
• Fixed: Potential access violation when Fakesheet mode was displaying note-based lyrics.
• Fixed: Staff window wasn't showing any bar-based lyrics.
• Fixed: 'Lyrics Below By' setting in Print Options changed to Bar Lyric Position so it will now let the user adjust the position of the bar lyrics whether they are displayed above or below.
• Fixed: Note colors now are working for any track, and will not be enabled when drum notation is in use.
• Fixed: When Song Title is blank, in Print Options, the Composer, Style, etc. would show up as garbage.
• Fixed: Copyright symbol would have an extra character in lead sheet if the copyright field was in use.

Summary of changes for DAW Plugin 3.6.15 (June 18, 2021)

• Fixed: StylePicker is showing 2020 as the newest styles instead of 2021.
• Fixed: More than one instance of BBM4.app would be running in some cases.
• Added: "Preview N Bars [Cmd/Ctrl-P]" option in Generate menu. Hot key for this function is CMD/CTRL-P
• Added: Option in "Preferences -> General Settings" to save current plugin size as default
• Added: Option in "Preferences -> Render Settings" to auto-clear renders on startup. Note that this option should only be enabled if the DAW creates copy of media when importing. Enabling this setting will delete the entire renders folder whenever the plugin is first loaded without checking if tracks were dragged or not.
• Added: Option in "Preferences -> Render Settings" to separate audio output channels for each track after generating
• Added: Notification for missing media (tracks that will not generate)
• Added: [PLAY] button to "Generation is complete!" message
• Added: AU plugin now sends MIDI to DLS Music Device if "Send MIDI to DAW" is selected
• Added: "Play Current Chord [Shift-Enter]" to right-click menu of chord sheet. Hot key for this function is SHIFT-ENTER
• Fixed: BB frequently hangs a long time on "Retrieving Style"
• Fixed: Changed "Paste Special" to "Paste Special (from Band-in-a-Box)" which is only enabled when system clipboard has song info
• Fixed: Plugin sometimes does not prompt to save generated tracks with song
• Fixed: Status message text has bad horizontal scale (should not be compressed)
• Fixed: Updated audio output channel display to show track channels -> output channels
• Fixed: "Chart" button text was cutoff
• Fixed: Play button in Chord Builder crashed AU plugin
• Fixed: "Preview Chord Progression" crashed AU plugin
• Fixed: Chordsheet should get focus when generation is complete
• Fixed: Issues with style picker (and other bb windows) hiding when CMD-TAB is used to navigate to another app and then back to DAW
• Fixed: Multiple instances of bbm4 open when running AU and VST plugin and retrieving style upon launch
• Fixed: Plugin should ask to save song before loading song demo
• Fixed: Preferences dialog should not move position when selected tab changes
• Fixed: Play/Stop buttons should still alert user that nothing is generated when they are disabled
• Fixed: Moved "Preview Chord Progression" feature from right-click menu to button under chord sheet (next to C7 button)

Summary of changes for build 503 (June 3, 2021)

• Fixed: Importing a 24 bit audio file would result in silence.
• Fixed: Recording audio with multi-channel interfaces might be silent.
• Fixed: "WMA" was displayed on the drop station instead of "AIFF".
• Fixed: Mono Waves AU plugins are silent.
• Fixed: Mono Waves VST plugins are silent on the right channel.
• Fixed: Recent song files in File menu might not load the correct song.
• Fixed: "Fix sour notes" default track data is not set properly for new songs.
• Fixed: File > Save Special > Save Song as M4A Audio creates corrupt file.
• Fixed: File > Save Special > Save Song as AIFF Audio creates corrupt file.
• Fixed: Copy Song to Text, long style name was showing a short name.
• Fixed: AddOns dialog link to webpage fixed.
• Fixed: Songs with held chord as final chord could crash.
• Added: Edit - Song Form - TIme Signature dialog now has settings for number of bars.

Summary of changes for DAW Plugin 3.5.20 (June 3, 2021)

• Fixed: Plugin sometimes crashes in Pro Tools (when setting negative value for bar offset)
• Fixed: Duplicate file name displayed in "Recent Files"

If you encounter a problem or have a concern with any of the features / functions of Band-in-a-Box^®, do not hesitate to contact us. Updated July 29, 2021

• • Added:
  • - support for NIKKOR Z 800mm f/6.3 VR S lenses,
  • - support for ML-L7 remote controls,
  • - Focus ring rotation range¹ to the CUSTOM SETTING MENU in position f9,
  • - Switch focus/control ring roles¹ to the CUSTOM SETTING MENU in position f10, and
  • - Save focus position and Recall focus position² to the roles that can be assigned using Custom Setting f2 (Custom controls) in the CUSTOM SETTING MENU.
• As of April 20, 2022, these features are available only with the following lenses:
• 1. NIKKOR Z 70-200mm f/2.8 VR S
• 2. NIKKOR Z 70-200mm f/2.8 VR S, NIKKOR Z 100-400mm f/4.5-5.6 VR S, and NIKKOR Z 400mm f/2.8 TC VR S
• • Made the following improvements:
  • - The camera focuses more readily on the subject in the center of the frame if it fails to detect the face or eyes of a portrait subject when Auto-area AF is selected for AF-area mode.
  • - Memory recall has been altered so that regardless of the focus mode selected, the focus position chosen via autofocus does not change if the shutter-release button is pressed halfway while memory recall is in effect.
• • Fixed the following issues:
  • - The camera would sometimes fail to focus on the subject in the focus area in pictures taken immediately after the battery was inserted.
  • - Reactivating the standby timer in User Settings mode U1, U2, or U3 would cause the shutter speed to change if the stored value was slower than 30 s.
  • - Pictures viewed by rotating the main command dial after zooming in on an image in “tall” (portrait) orientation in the viewfinder would not display correctly.
  • - Jitter would occur in movies recorded while footage was being output to an HDMI device with 3840×2160; 60p or 3840×2160; 50p selected for Frame size/frame rate in the MOVIE SHOOTING MENU.
  • - The movie-record button would not function with certain gimbals while video was being recorded to an external device via HDMI with 3840×2160; 30p, 3840×2160; 25p, or 3840×2160; 24p selected for Frame size/frame rate in the MOVIE SHOOTING MENU and 10 bit selected for Advanced > Output data depth in the SETUP MENU.

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Fixed editing workspace for Graph Editor toolbar when it was not visible. Zippyshare. Proplanner will then determine your eligibility for an extended license and will email you an activation key code that you put in the same window as Figure B. Uninstall LicenseServer and install it. I am unable to tick the "loaded" box in plugin manager(the tick would disappear once my mouse leave the box). Click Next. act file to upload, (this file is different than the Continuum123456. Nrf2 activation assay 3251a877d4 Renderman Pro Server 17 Crackl · J. 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The customer portal contains the ability to view licenses and servers associated with your Step 7: Copy and paste your Sapphire Activation Key license that you received from your license email into the “License activation key” field. Download All Files. Choose [Add/Renew Licenses]. With annual subscription for Modo, you can get easy, affordable access to the latest version of Modo. 1 998 1 GreyscaleGorilla Light Kit Pro v3 for Cinema 4D R18 to R20 Win/Mac 223 / 817 MB Read More. Download Boris FX Full Version – BorisFX Sapphire is a 3rd party plugin that has various cool effects features for Adobe and OFX. The number after the “=” is the timeout time. Click Activate. (1987). Step 4: License file to create or edit: name it something like vm_enable. Mar 16, 2021 — Boris FX Sapphire the previous name of the company: Genarts is a collection of powerful plugins for creating visual effects and effects in movies . 2 Crack + Serial Key Free Download Full Version. Platform: All Platforms. 31. Activation Pro gives you, the software publisher, the ability to deliver electronic licenses to your customers 24x7, without customer support involvement. 1 This web interface is used to administer the RLM Activation Pro database . 0. Terrestrial laser scanning (TLS) registration. The lab admin should check that the keyserver software has been installed; that it is running; and that the host computer is online. Go to [License Management] menu. Version 10: Install the program and before running, run the rlm file located in the path Crack \ statease-rlm with Run as administrator. a. ----- If you skipped the license installation during the initial installation process, you can follow the instructions below: On Windows 7, go to: Start > All Programs > GenArts Sapphire AE. This software, which offers a set of functional features, is compatible with the most popular CG software and is suitable for use in studios or by creative artists

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• • Added:
  • - support for NIKKOR Z 800mm f/6.3 VR S lenses,
  • - support for ML-L7 remote controls,
  • - Focus ring rotation range¹ to the CUSTOM SETTING MENU in position f9,
  • - Switch focus/control ring roles¹ to the CUSTOM SETTING MENU in position f10, and
  • - Save focus position and Recall focus position² form z mac Activators Patch the roles that can be assigned using Custom Setting f2 (Custom controls) in the CUSTOM SETTING MENU.
• As of April 20, 2022, these features are available only with the following lenses:
• 1. NIKKOR Z 70-200mm f/2.8 VR S
• 2. NIKKOR Z 70-200mm f/2.8 VR S, NIKKOR Z 100-400mm f/4.5-5.6 VR S, and NIKKOR Z 400mm f/2.8 TC VR S
• • Made the following improvements:
  • - The camera focuses more readily on the subject in the center of the frame if it fails to detect the face or eyes of a portrait subject when Auto-area AF is selected for AF-area mode.
  • - Memory recall has been form z mac Activators Patch so that regardless of the focus mode selected, the focus position chosen via autofocus does not change if the shutter-release button magix vegas movie studio platinum 17 pressed halfway while memory recall is in effect.
• • Fixed the following issues:
  • - The camera would sometimes fail to focus on the subject in the focus area in pictures taken immediately after the battery was inserted.
  • - Reactivating the standby timer in User Settings mode U1, U2, or U3 would cause the shutter speed to change if the stored value was slower than 30 s.
  • - Pictures viewed by rotating the main command dial after zooming in on an image in “tall” (portrait) orientation in the viewfinder would not display correctly.
  • - Jitter would occur in movies recorded while footage was being output to an HDMI device with 3840×2160; 60p or 3840×2160; 50p selected for Frame size/frame rate in the MOVIE SHOOTING MENU.
  • - The movie-record button would not form z mac Activators Patch with certain gimbals while video was being recorded to an external device via HDMI with 3840×2160; 30p, 3840×2160; 25p, or 3840×2160; 24p selected for Frame size/frame rate in the MOVIE SHOOTING MENU and 10 bit selected for Advanced > Output data depth in the SETUP MENU.

MACINTOSH:

WINDOWS:

Band-in-a-Box^® 2021 for Mac Update - Build 511 (100 MB)

This patch will update Band-in-a-Box^® 2021 to build 511 from any previous 2021 build. This includes DAW Plugin 3.7.23. You need to have Band-in-a-Box^® 2021 (build 502 or higher) installed for this update to work.

If you are updating an external PG Music hard drive, make sure to choose the correct destination volume ([Change Install Location] button / Install on a specific disk. NOTE: if you get a message that it must be installed on the system drive, close other running applications and start the installer again, or if that does not resolve it, reboot the Mac.

Note: If you have a localized version (German, French, Italian), you should not install this patch, but should instead download the latest patch from here.

Summary of changes for build 511 (July 29, 2021)

• Fixed: The MIDI keyboard wizard might not work during recording.
• Fixed: "flat, dry, and center" option in the Drop Plus dialog should be available for dragging single tracks.
• Fixed: MIDI files containing sysex data would fail to import.
• Fixed: Dropping a MIDI file onto Band-in-a-Box would be ignored if the file extension was ".midi".
• Fixed: Mouse wheel scrolling did not work in the MIDI Import dialog.
• Fixed: When importing MIDI files "Merge with existing data" did not work correctly.
• Fixed: Undoing audio edits on tracks Utility#2 or higher did not work.
• Fixed: Cut/Copy/Paste hot keys did not work in Open/Save dialogs if launched from debut video capture registration code 2019 Free Activators modal dialog (eg the Render dialog).
• Fixed: When asked to overwrite song file [No] should be the default button (activated form z mac Activators Patch Return key).
• Fixed: The [Cancel] button in popup dialogs should selectable with tab and arrow keys (activated by Space Bar).
• Fixed: Erase Track causes tracks to change solo state.

Summary of changes for DAW Plugin 3.7.23 (July 29, 2021)

• Fixed: VST3 did not correctly load the dedicated synth
• Fixed: "Set default style" not working
• Fixed: Moving files when saving doesn't work if style is loaded but not generated
• Fixed: DLS Music Device not loading in VST3.
• Added: Preview N Bars to chord sheet right-click menu
• Added: Message notifying that saved window size has completed
• Added: Bar highlighting for midi preview
• Fixed: Clicking anywhere on plugin while bbm4 window is open should bring window to front (plugin will now be greyed out while bb dialog is open)
• Fixed: Change disabled Preview N Bars to Preview 4 Bars by default
• Fixed: Chord sheet/track table view should not resize if horizontal size has been changed
• Fixed: Some strings not externalized in chord builder
• Fixed: PLAY/STOP buttons should be green/black to match BIAB
• Fixed: Gain value changes when toggling volume slider
• Fixed: "Move media to project directory" when saving grammarly premium Crack Key For U not always working
• Fixed: Limit for choruses and start bar were incorrect
• Fixed: Selecting secondary chord display and then going back to none doesn't resize rows well
• Fixed: Components incorrectly form z mac Activators Patch in confirmation dialog
• Fixed: Expand clear renders feature to exclude currently generated tracks
• Fixed: Expand auto-clear feature to work when starting new session
• Fixed: Plugin sometimes resets when closing/reopening plugin editor
• Fixed: "Auto-generate song" was not working with song demos
• Fixed: noise when starting sync playback in the plugin
• Fixed: chord sheet should get keyboard focus after any action
• Fixed: generate "region" not working
• Fixed: Save Window Size was not working

Summary of changes for build 509 (July 08, 2021)

• Fixed: Plugins not saved correctly in song files.
• Fixed: Frozen RealDrums RealCharts were not saved in song files.
• Fixed: Issues with different tempo % changes at different choruses, form z mac Activators Patch, and starting playback in different choruses.

Summary of changes for build 507

• Fixed: The time signature does not update on the chord form z mac Activators Patch after selecting a style.
• Fixed: Using mouse wheel over Track Selector control does not work to select None (eg in Record dialog).
• Fixed: Drag and drop Master track to MIDI quadrant, and 1 track is soloed, should be SMF type 0 instead of 1.
• Fixed: Band-in-a-Box should automatically copy Factory_stylz1.in to stylz1.bin if stylz1.bin is missing.
• Fixed: File > Revert to saved should not ask user if they want to save their current song.
• Fixed: Issues with chord sheet scrolling when near end of song.
• Fixed: Cancel should be triggered with [Esc] key in the post form z mac Activators Patch audio dialog.
• Fixed: Plugins on Utility tracks do not save in song files.
• Fixed: Track Actions > Save as performance track would fail.
• Fixed: Using Copy/Move Tracks to copy audio to a non-editable track (eg Soloist) would fail.
• Fixed: Possible crash when opening the Notation window.
• Fixed: When choosing a different language the message "New Language will take effect the next time you run the program." should be displayed in the new language as well as the current language.
• Fixed: Key sig being drawn with the wrong font and position.
• Fixed: Drum notes in bottom staff that were saved to musicXML would display way too low when loaded into other programs.
• Added: Feature Browser menu item added to the Help menu.

Summary of changes for build 506 (June 29, 2021)

• Added: "Avoid connecting to low quality input devices" setting added to Audio Devices dialog.
• Fixed: Audio Unit plugins (eg the default synth) might be silent with some Audio Devices (eg bluetooth headphones).
• Fixed: Delete Beats at Bar was not undoable.
• Fixed: Delete Beats at Bar should ask for chorus/bar numbers rather than just a bar number.
• Fixed: Insert Beats at Bar should ask for chorus/bar numbers rather than just a bar number.
• Fixed: Chords and bar lines might not line up properly in the Piano Roll.
• Fixed: "Utility Tracks were automatically transposed." message shows after leaving song settings when no transpose settings were changed.

Summary of changes for build 505

• Fixed: Memory leaks when regenerating RealTracks or playing songs several times in a program session.
• Fixed: Styles that contain loops might try to load a WMA file and crash the program.
• Fixed: Quick render menu items should only render the song mix, and ignore "One file per track" setting.
• Fixed: Quick render menu items were ignoring the "Save in current song directory" setting.
• Fixed: Track context menu > Track Actions > Save track as WAV file should select stereo/mono automatically.
• Fixed: "Also generate MIDI Drums in the MIDI file" did not export the underlying MIDI style if the song has RealDrums RealCharts.
• Fixed: New Xtra Style might not display correctly in the StylePicker.
• Fixed: If scanning or loading a VST plugin causes Band-in-a-Box to crash, then that plugin should be automatically excluded the next time running the program.
• Added: Plugin selector dialog shows info on listed plugins, such as the file path and unique identifier. This is useful for troubleshooting plugin installation problems, duplicates, etc.
• Added: "Allow multiple instances of Band-in-a-Box to be running" setting added to the Preferences dialog.
• Added: [MIDI file] button added to the Preferences dialog. This replaces all of the check boxes related to MIDI exporting.
• Added: Batch Mode for exporting MIDI files is now accessible from the MIDI export dialog.
• Improved: Audio Devices dialog is resizable.

• Fixed: Partially hidden check boxes in the MIDI File Options dialog.
• Fixed: Batch Mode button should not be showing in the MIDI export dialog.
• Fixed: Drag and drop render might cause an hour glass mouse cursor to get stuck.
• Fixed: Drag and drop individual MIDI tracks form z mac Activators Patch be SMF type 0 instead of 1.
• Fixed: Frozen RealTracks on Drums track are not eset smart security premium 11 Crack Key For U in song.
• Fixed: VST plugin scans may be slower in some cases.
• Fixed: It was not possible to select audio inputs on multi-channel audio interfaces.
• Fixed: New Xtra styles do not show mobaxterm professional 11 crack Free Activators the StylePicker for localized versions if the language patch is not up to date.
• Improved: File rendering time reduced.
• Improved: Enabling render setting "Individual tracks flat, dry, and center" will improve rendering time.
• Added: "Include Mix" option added to rendering options which can be disabled if rendering individual tracks.
• Added: "Save in current song directory" setting added to Render dialog.
• Added: MIDI Chord Detection window added. This can be accessed from Window > MIDI Chord Detection.
• Fixed: Potential access violation when Fakesheet mode was displaying note-based lyrics.
• Fixed: Staff window wasn't showing any bar-based lyrics.
• Fixed: 'Lyrics Below By' setting in Print Options changed to Bar Lyric Position so it will now let the user adjust the position of the bar lyrics whether they are displayed above or below.
• Fixed: Note colors now are working for any track, and will not be enabled when drum notation is in use.
• Fixed: When Song Title is blank, in Print Options, the Composer, Style, etc. would show up as garbage.
• Fixed: Copyright symbol would have an extra character in lead sheet if the copyright field was in use.

Summary of changes for DAW Plugin 3.6.15 (June 18, 2021)

• Fixed: StylePicker is showing 2020 as the newest styles instead of 2021.
• Fixed: More than one instance of BBM4.app would be running in some cases.
• Added: "Preview N Bars [Cmd/Ctrl-P]" option in Generate menu, form z mac Activators Patch. Hot key for this function is CMD/CTRL-P
• Added: Option in "Preferences -> General Settings" to save current plugin size as default
• Added: Option in "Preferences -> Render Settings" to auto-clear renders on startup. Note that this option should only be enabled if the DAW creates copy of media when importing. Enabling this setting will delete the entire renders folder whenever the plugin is first loaded without checking if tracks were dragged or not.
• Added: Option in "Preferences -> Render Settings" to separate audio output channels for each track after generating
• Added: Notification for missing media (tracks that will not generate)
• Added: [PLAY] button to "Generation is complete!" message
• Added: AU plugin now sends MIDI to DLS Music Device if "Send MIDI to DAW" is selected
• Added: "Play Current Chord [Shift-Enter]" to right-click menu of chord sheet. Hot key for this function is SHIFT-ENTER
• Fixed: Form z mac Activators Patch frequently hangs a long time on "Retrieving Style"
• Fixed: Changed "Paste Special" to "Paste Special (from Band-in-a-Box)" which is only enabled when system clipboard has song info
• Fixed: Plugin sometimes does not prompt to save generated tracks with song
• Fixed: Status message text has bad horizontal scale (should not be compressed)
• Fixed: Updated audio output channel display to show track channels -> output channels
• Fixed: "Chart" button text was cutoff
• Fixed: Play button in Chord Builder form z mac Activators Patch AU plugin
• Fixed: "Preview Chord Progression" crashed AU plugin
• Fixed: Chordsheet should get focus when generation is complete
• Fixed: Issues with style picker (and other bb windows) hiding when CMD-TAB is used to navigate to another app and then back to DAW
• Fixed: Multiple instances of bbm4 open when running AU and VST plugin and retrieving style upon launch
• Fixed: Plugin should ask to save song before loading song demo
• Fixed: Preferences dialog should not move position when selected tab changes
• Fixed: Play/Stop buttons should still alert user that nothing is generated when they are disabled
• Fixed: Moved "Preview Chord Progression" feature from right-click menu to button under chord sheet (next to C7 button)

Summary of changes for build 503 (June 3, 2021)

• Fixed: Importing a 24 bit audio file would result in silence.
• Fixed: Recording audio with multi-channel interfaces might be silent.
• Fixed: "WMA" was displayed on the drop station instead of "AIFF".
• Fixed: Mono Waves AU plugins are silent.
• Fixed: Mono Waves VST plugins are silent on the right channel.
• Fixed: Recent song files in File menu might not load the correct song.
• Fixed: "Fix sour notes" default track data is not set properly for new songs.
• Fixed: File > Save Special > Save Song as M4A Audio creates corrupt file.
• Fixed: File > Save Special > Save Song as AIFF Audio creates corrupt file.
• Fixed: Copy Song to Text, long style name was showing a short name.
• Fixed: AddOns dialog link to webpage fixed.
• Fixed: Songs with held chord as final chord could crash.
• Added: Edit - Song Form - TIme Signature dialog now has settings for number of bars.

Summary of changes for DAW Plugin 3.5.20 (June 3, 2021)

• Fixed: Plugin sometimes crashes in Pro Tools (when setting negative value for bar offset)
• Fixed: Duplicate file name displayed in "Recent Files"

If you encounter a problem or have a concern with any of the features / functions of Band-in-a-Box^®, do not hesitate to contact us. Updated July 29, 2021

Sublytic Membrane-Attack-Complex (MAC) Activation Alters Regulated Rather than Constitutive Vascular Endothelial Growth Factor (VEGF) Secretion in Retinal Pigment Epithelium Monolayers^*

Kannan Kunchithapautham and Bärbel Rohrer¹

Kannan Kunchithapautham

From the Departments of Ophthalmology and Neurosciences Division of Research, Medical University of South Carolina, Charleston, South Carolina 29425

Find articles by Kannan Kunchithapautham

Bärbel Rohrer

From the Departments of Ophthalmology and Neurosciences Division of Research, Medical University of South Carolina, Charleston, South Carolina 29425

Find articles by Bärbel Rohrer

Author informationArticle notesCopyright and License informationDisclaimer

From the Departments of Ophthalmology and Neurosciences Division of Research, Medical University of South Carolina, Charleston, South Carolina 29425

¹ A Research to Prevent Blindness Olga Keith Weiss Scholar. To whom correspondence should be addressed: Department of Ophthalmology, Medical University of South Carolina, 167 Ashley Ave., SEI 511, Charleston, SC, 29425., Tel.: 843-792-5086; Fax: 843-792-1723; E-mail: ude.csum@rerhor.

Received 2010 Dec 21; Revised 2011 Apr 27

Copyright © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

Abstract

Uncontrolled activation of the alternative complement pathway and secretion of vascular endothelial growth factor (VEGF) are thought to be associated with age-related macular degeneration (AMD). Previously, we have shown that in RPE monolayers, oxidative-stress reduced complement inhibition on the cell surface. The resulting increased level of sublytic complement activation resulted in VEGF release, which disrupted the barrier facility of these cells as determined by transepithelial resistance (TER) measurements. Induced rather than basal VEGF release in RPE is thought to be controlled by different mechanisms, including voltage-dependent calcium channel (VDCC) activation and mitogen-activated protein kinases. Here we examined the potential intracellular links between sublytic complement activation and VEGF release in RPE cells challenged with H₂O₂ and complement-sufficient normal human serum (NHS). Disruption of barrier function by H₂O₂ + NHS rapidly increased Ras expression and Erk and Src phosphorylation, but had no effect on P38 phosphorylation. Either treatment alone had little effect. TER reduction could be attenuated by inhibiting Ras, Erk and Src activation, or blocking VDCC or VEGF-R2 activation, but not by inhibiting P38. Combinatorial analysis of inhibitor effects demonstrated that sublytic complement activation triggers VEGF secretion via two pathways, Src and Ras-Erk, with the latter being amplified by VEGF-R2 activation, but has no effect on constitutive VEGF secretion mediated via P38. Finally, effects on TER were directly correlated with release of VEGF; and sublytic MAC activation decreased levels of zfp36, a negative modulator of VEGF transcription, resulting in increased VEGF expression. Taken together, identifying how sublytic MAC induces VEGF expression and secretion might offer opportunities to selectively inhibit pathological VEGF release only.

Keywords: Aging, Complement, Growth Factors, Oxidative Stress, Signal Transduction, Alternative Pathway, Angiogenesis, Retinal Pigment Epithelium, Transepithelial Resistance, Vascular Endothelial Growth Factor

Introduction

Age-related macular degeneration (AMD),² the leading cause of blindness for Americans over age sixty, is a slowly progressing multifactorial disease involving genetic abnormalities and environmental insults. Whereas mechanistic studies have shown that inflammation (1) and smoking (2) are fundamental components of AMD, genetic studies have demonstrated that polymorphisms in different complement proteins each increase the risk for developing AMD. Genetic variations with a number of complement components, including those of required activators (complement factor B, CFB) (3), inhibitors (complement factor H, CFH (4–6); complement 1 inhibitor, C1INH, although the evidence is conflicting (7, 8)) and essential components in the complement cascade (complement factor 2 and 3, C2 (3) and C3 (9)) have been found to be associated with all forms of AMD. In particular, one of the most detrimental mutations occurs in CFH (fH risk haplotype). The only environmental agent unequivocally linked to AMD is nicotine (10), presumably by generating oxidative stress. Overall, oxidative stress is thought to be an important risk factor, presumably impacting RPE function (11). Current available treatments for AMD focus on the late stage of the disease (choroidal neovascularization; CNV). To this end, therapies have focused on either physically or pharmacologically interrupting blood vessel growth (summarized in the following review article (12)), in particular, by using anti-angiogenic drugs (VEGF inhibitors) that deplete the new blood vessels of an essential growth factor. Finally, what all stages of AMD have in common is varying degrees of damage to the RPE/Bruch's membrane, which ultimately results in damage to the underlying photoreceptors.

Early AMD is characterized by the formation of drusen at the choroid/RPE interface (1, 13–15). Drusen contain large numbers of proteins including amyloid proteins, coagulation factors and complement components. A high concentration of complement regulatory proteins and membrane attack complex (MAC), in the area of Bruch's membrane and the RPE has been documented (1, 15–17). MAC deposition density is correlated with severity of AMD and is most pronounced in the macula. Furthermore, there is a correlation between the amount of MAC deposition and the loss of RPE cells (1). These data, combined with the relative lack of immunoglobulins in drusen (which would activate the classical pathway) support the hypothesis that the alternative pathway may play a critical role in driving the pathology of AMD. Thus, the concept emerges that abnormalities in controlling the AP may lead to inflammation at the level of the RPE/Bruch's membrane, generating a pathological environment that includes drusen formation and VEGF production and secretion, which is favorable for the development of AMD.

To further examine this concept, we investigated RPE cells (ARPE-19 cells) grown as stable monolayers (18). These monolayers exhibit stable transepithelial resistance of ∼40–45 ohm/cm²; are polarized; and stain for markers of tight and adherence junctions (19). In addition, the Cheetham group has shown that ARPE-19 cells when grown as monolayers, are resistant to oxidative damage; while subconfluent ARPE-19 cells exposed to oxidative stress (≥0.1 mm H₂O₂) undergo apoptosis (20), those grown in monolayers show no sign of damage up to 0.5 mm (19). The earliest sign of damage occurs with H₂O₂ concentrations above 1 mm, resulting in a reduction of transepithelial resistance (TER) (19). Thus, we asked whether a concentration that did not affect the epithelial integrity (0.1 mm H₂O₂), sensitized the cells to complement attack. Based on these experiments (18), we proposed a dual-hit hypothesis in RPE damage, generating a conducive environment for AMD pathology. Oxidative stress in RPE was found to decrease the levels of alternative pathway inhibition by reducing surface expression of the complement inhibitors, CD55 and CD59, and by impairing alternative pathway regulation at the cell surface by factor H present within the serum. Together, these changes sensitized the RPE to complement attack, resulting in increased VEGF secretion and TER reduction.

Here we extend these analyses by investigating how sublytic MAC might be controlling VEGF release and expression. Sublytic MAC-activation results in the formation of discrete channels consisting of a tetramolecular C5b-8 and the tubular poly-C9 complex, which allows for the influx of mainly Na⁺ and Ca²⁺ ions. Local influx of Ca²⁺ might directly control VEGF secretion. However, other pathways should be considered. The Strauss' laboratory has shown that VEGF secretion can be controlled by the activity of voltage-gated calcium channels (VDCC), which themselves are regulated by Src kinase (21); whereas work by Klettner and Roider has shown that VEGF secretion from RPE cells is controlled by two pathways (P38 MAP kinase controls constitutive secretion, whereas Erk kinase signaling controls stimulated secretion (22)). The results presented here demonstrate that sublytic MAC utilizes the regulated VEGF secretory pathway to increase VEGF release and production. These results are discussed in the context of VEGF blocking strategies that interfere with both basal VEGF levels required for the health of ocular tissues and the pathological VEGF production seen in ocular inflammatory diseases.

EXPERIMENTAL PROCEDURES

Reagents

The reagents used in these studies included pooled normal human serum (NHS (Quidel)) as a source of complement proteins. Primary antibodies to phospho-P38, P38; phospho-Erk, Erk; phospho-Src, Src; and GAPDH were purchased from Cell Signaling Technology, Inc. (Danvers, MA); the primary antibody to Ras was obtained from Millipore (Billerica, MA). To analyze phosphoproteins, a protease inhibitor mixture was added (Sigma Aldrich). Species-specific secondary antibodies were from Jackson ImmunoResearch (West Grove, PA) and MP Biomedicals, Inc. (Santa Ana, CA). The VEGFR-1/2 inhibitor, SU5416 (Chemicon; now Millipore), was used to block the effects of VEGF. SU5416 (Z-3-[(2,4-dimethylpyrrol-5-yl) methylidenyl]-2-indolinone) is a lipophilic synthetic receptor tyrosine kinase inhibitor, which inhibits VEGFR-1/2 by binding to the ATP-binding pocket within the kinase domain of the receptor. SU5416 has been shown to inhibit VEGF-dependent endothelial cell proliferation in vitro and in animal models. Nifedipine (TOCRIS Bioscience, Ellisville, MI) is a dihydropyridine or L-type calcium channel blocker. It was used to stabilize the VDCCs in the closed position as described in primary human RPE cells (21). Kinase inhibition was achieved using U0126 (1,4-diamino-2,3-dicyano-1,4-bis(2-amino-phenylthio)butadiene; Promega), a highly selective inhibitor of both MEK1 and MEK2 (23); FTS (S-trans, transfarnesylthiosalicylic acid; Cayman Chemical Co., Ann Arbor, MI), a non-toxic but selective Ras antagonist (24); PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyraz) lo[3,4-d]pyrimidine; Calbiochem), a potent and reversible, ATP-competitive inhibitor of c-Src (25); as well as SB203580 (4-[4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-1H-imidazol-5-yl]pyridine; Promega), a potent pyridinyl imidazole inhibitor of P38 MAP kinase (26).

Cell Culture System

Experiments were performed using ARPE-19 cells, a human retinal pigment epithelial cell line that displays the differentiated phenotype of RPE cells, and forms a polarized monolayer on Transwell filters (Costar; (27, 28)). Cells were grown in DMEM-F12 (Invitrogen) with 10% fetal bovine serum (FBS) and 1× penicillin:streptomycin as monolayers on Transwell filters. FBS was removed completely for the final 5–7 days (2–3 media changes) prior to measurements, which we have previously shown does not alter survival or monolayer formation in these cells (29). TER of the cell monolayer on the Transwell filters was determined by measuring the resistance across the monolayer with an EVOM volt-ohmmeter (World Precision Instruments) (18). The value for cell monolayers was determined by subtracting the TER for filters without cells and then multiplying by the surface area of the filters. Cell monolayers were considered stable when TER was repeatedly measured as ∼40–45 Ω/cm² (29). TER measurements, which are proportional to membrane permeability, are an accepted readout for the barrier function of an RPE monolayer (27, 29).

As a model of oxidative stress, stable ARPE-19 cell monolayers were treated with 0.5 mm of H₂O₂. It has previously been reported that doses of up to 1 mm are not cytotoxic, and do not lead to disruption of barrier function in these cells (19). After treatment with H₂O₂, monolayers were immediately exposed to 25% NHS as a source of complement proteins (18).

Quantitative RT-PCR

ARPE-19 cells from control and experimental sets were collected and stored at 80 °C until used. Quantitative RT-PCR analyses were performed as described in detail previously (30). Primers used were: β-actin, forward: 5′-AAATCTGGCACCACACCTTC-3′ and reverse: 5′-GGGGTGTTGAAGGTCTCAAA-3′; VEGF forward: 5′-AAGGAGGAGGCAGAATCAT-3′ and reverse: 5′-CACACAGGATGGCTTGAAGA-3′; zfp36 forward: 5′-ATCGCCACCCCAAATACAAG-3′ and reverse: 5′-GTCTTCGCTAGGGTTGTGGA-3′. Real-time PCR analyses were performed in triplicate in a GeneAmp® 5700 Sequence Detection System (Applied Biosystems, Foster City, CA) using standard cycling conditions. Quantitative values were obtained by the cycle number.

ELISA for VEGF

To measure production of VEGF by ARPE-19 cells, apical and basal supernatants were concentrated (Amicon Ultra-4, 3000 Da cutoff; Millipore), solubilized in CellLytic MT (mammalian tissue lysis/extraction reagent; Sigma) and centrifuged at 20,000 × g for 5 min. Microplates were coated with the anti-human VEGF polyclonal capture antibody (Antigenix America, Inc.; Huntington Station, NY) (18), and 100 μl of supernatant was added. The captured proteins were detected with the same VEGF-specific antibody conjugated to horseradish peroxidase (HRP), followed by development with the chromogenic substrate, OPD (Sigma Aldrich). Product development was assayed by measuring absorbance at 492 nm. Aliquots were assayed in duplicate, and values compared with a VEGF dose-response curve.

Western Blot Analysis

Cell extracts were separated by electrophoresis on a 10% Bis-Tris polyacrylamide gel (Invitrogen, Carlsbad, CA), and proteins were transferred to a nitrocellulose membrane. The membrane was probed with primary antibodies; antibody binding was visualized using a chemiluminescence detection kit (Immobilon Western; Millipore Corporation, Billerica, MA). The intensity of the bands was quantified using the Alpha Innotech Fluorchem 9900 imaging system running Alpha Ease FC Software 3.3 (Alpha Innotech, San Leandro, CA). As a loading control, blots were stripped and reprobed with an antibody against GAPDH (Stressgen, Ann Arbor, MI).

Statistics

For data consisting of multiple groups, one-way ANOVA followed by Fisher's post hoc test (p < 0.05) was used; single comparisons were analyzed by t test analysis (p < 0.05).

RESULTS

Sublytic MAC Activation Leads to Ras, Erk, and Src Activation

Sublytic complement activation, which is known to result in Ca²⁺ influx through the pores generated by the MAC, is associated with an increase in VEGF secretion (18). Here we examined whether the known kinase pathways involved in VEGF secretion, are activated by sublytic MAC activation. Kinase activation in ARPE-19 cell monolayers was investigated by Western blot analysis at baseline and within a 1-h time course after the exposure of monolayers to 0.5 mm H₂O₂ + 25% complement-sufficient normal human serum (NHS) (Fig. 1A). In particular we focused on P38 MAP kinase, which participates in constitutive VEGF secretion (22); Erk kinase, which participates in regulated VEGF secretion (22); Src, which increases VDCC-mediated VEGF secretion (21); and finally Ras, which provides a link between Src and Erk signaling.

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FIGURE 1.

Complement activation on ARPE-19 cells induces intracellular signaling associated with regulated VEGF secretion. ARPE-19 cells were grown as a monolayer until a stable TER was obtained, at which point serum was withdrawn for 5–7 days prior to the experiment. A, following apical treatment with 0.5 mm H₂O₂ and 25% NHS for the specified times, cells were collected and analyzed for protein expression by Western blotting. Complement activation rapidly and transiently induced Erk phosphorylation and Ras expression; rapidly and sustainably induced Src-phosphorylation; but partially suppressed P38 phosphorylation. B, controls were examined at the 5-min time point, demonstrating that 0.5 mm H₂O₂ or 25% NHS, each administered alone, had no effect on expression levels and phosphorylation status of the signaling molecules. C, summary of expression levels and phosphorylation status of the signaling molecules. Data are expressed as mean ± S.E. (n = 3 per condition).

P38 phosphorylation was found to be reduced by ∼40% after the exposure of monolayers to H₂O₂ + NHS, when compared with the overall amount of P38 present in the monolayers (Fig. 1, A and C), and remained depressed during the time course analyzed (1 h). A transient increase in phospho-Erk could be detected rapidly within minutes, as could overall Ras levels (Fig. 1, A and C). Both Erk and Ras activation peaked within 5 min after the onset of stimulation (earliest time point analyzed) and returned to baseline levels by 10 (Ras) or 20 min (Erk) (Fig. 1, A and C). In contrast, while the increase in phosphorylation of Src is equally rapid, it remained elevated over the 1-h investigation period (Fig. 1, A and C). H₂O₂ or NHS when administered singly did not lead to any significant alterations in kinase activity (Fig. 1B).

Sublytic MAC-mediated TER Reduction Is Sensitive to Ras, Erk, Src, and VDCC Inhibition

Sublytic MAC formation is associated with an increase in VEGF secretion, leading to TER reduction, which can be inhibited by using either an alternative pathway inhibitor or by blocking VEGF receptor-1/2 signaling (18). Thus, if the above-investigated pathways were to be involved in VEGF secretion, inhibiting their activation should blunt TER reduction produced by H₂O₂ + NHS (Fig. 2A) and reduce VEGF secretion (Fig. 2B).

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FIGURE 2.

Disruption of barrier function by complement activation on ARPE-19 cells is correlated with activity in pathways known to control regulated secretion of VEGF. Serum-deprived ARPE-19 cell monolayers with stable TER were treated with 0.5 mm H₂O₂ and 25% NHS in the presence and absence of pathway-specific inhibitors. A, treatment with H₂O₂ and serum caused a reduction in transepithelial resistance (TER) by ∼40% after 4 h. Co-administration of nifedipine (a dihydropyridine or L-type calcium channel blocker); U0126 (a highly selective inhibitor of Erk); FTS (a selective Ras antagonist); or PP2 (a reversible, ATP-competitive inhibitor of c-Src) blunted the deterioration of TER induced by H₂O₂ + 25% NHS; whereas SB203580 (a potent inhibitor of P38 MAP kinase) had no effect. B, after TER measurements, apical supernatants were removed and analyzed for VEGF content. VEGF secretion is not altered by H₂O₂ or NHS, whereas H₂O₂ + NHS together resulted in a significant increase in VEGF release. The inhibitors that blunted TER reduction also resulted in a reduction in VEGF secretion. C, when VEGF secretion was plotted against the percentage reduction in TER, a perfect correlation was revealed. Data are expressed as mean ± S.E. (n = 3 per condition).

As shown previously, TER is stable over the 4-hour recording period in control monolayers, but is reduced by ∼50% in those treated with H₂O₂ + NHS (Fig. 2A). Adding 10 μm nifedipine, a specific inhibitor for VDCC (21), to the monolayers exposed to H₂O₂ + NHS almost completely prevented the deterioration of TER. Likewise, the effect of H₂O₂ + NHS is blunted when inhibiting Erk kinase with 10 μm U0126 (23); Ras with 10 μm FTS (24); or Src with 4 pm PP2 (25). Adding the P38 MAP kinase inhibitor, SB203580 at 10 μm (26), had no effect on TER in the presence of H₂O₂ + NHS. None of the inhibitors had any effect on TER in the absence of H₂O₂ + NHS (data not shown).

The effects of the kinase and the VDCC inhibitor on MAC-induced TER reduction were found to be due to their effects on the secretion of VEGF, rather than other nonspecific effects (Fig. 2B). We have shown previously that co-administering H₂O₂ + NHS to either the apical or basal surface resulted in a large fold-increase in VEGF secretion, which was biased toward the apical surface (18). Here we confirmed by using VEGF ELISA assays that VEGF secretion from the RPE monolayers into the apical compartment was induced by H₂O₂ + NHS, whereas H₂O₂ or NHS had no significant effect. While the four inhibitors (nifedipine, U0126, FTS, and PP2) that were found to ameliorate the MAC-induced TER reduction, also significantly blunted the release of VEGF, the P38 MAP kinase inhibitor neither blunted TER reduction induced by H₂O₂ + NHS nor did it decrease VEGF secretion.

Sublytic MAC-mediated Intracellular Cross-talk

The analysis thus far has confirmed that sublytic MAC activation results in VEGF-mediated TER reduction; and that activation can be inhibited by blocking Ras, Erk, Src, and VDCC activity. However, it is unclear whether sublytic MAC triggers these pathways independently, or whether there is any cross-talk. In addition, it needs to be kept in mind that VEGF, which is secreted into the media in response to sublytic MAC-activation, will activate VEGF-R2 receptors that also signal through some of these molecules. To further characterize the intracellular signaling pathways, Western blotting to examine phosphorylation of Erk and Src, and levels of Ras expression were performed (Fig. 3A) and quantified (Fig. 3B) on lysates from cells pretreated with Erk, Src, Ras, VEGF-R2, and VDCC inhibitors, and then exposed to H₂O₂ + NHS for 5 min. As indicated before, H₂O₂ + NHS lead to the increase in Erk, Src, and Ras activation. The Erk inhibitor, U0126, only eliminated Erk activation; the Src inhibitor, PP2, eliminated Src activation and also reduced Ras levels; the Ras inhibitor, FTS, eliminated the increase in Ras expression, completely eliminated Erk, and slightly reduced Src phosphorylation; inhibiting VEGF-R2 with SU5416 blunted Src phosphorylation; and finally the VDCC inhibitor, nifedipine, reduced Src phosphorylation and Ras expression. This analysis places Ras upstream of Erk, but indicates that there is also cross-talk between Src and Ras. Since VEGF-R2 activation was found to stimulate Src phosphorylation, it might therefore amplify secretion of its own ligand. And finally, the effects of nifedipine, resulting in a reduction of Src phosphorylation and Ras expression levels, might be due to the fact that inhibiting the VDCC reduced VEGF secretion may have in part prevented activation of the VEGF-R2-dependent kinase activity.

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FIGURE 3.

Analysis of cross-talk between signaling pathways induced by complement activation in ARPE-19 cells. ARPE-19 cell monolayers were grown as described in Fig. 1. Erk and Src phosphorylation as well as Ras expression were analyzed in the presence of pathway-specific inhibitors. A, following apical treatment with 0.5 mm H₂O₂ and 25% NHS for 5 min, cells were collected and analyzed for protein expression by Western blotting. Complement activation rapidly induced Erk and Src phosphorylation as well as Ras expression. The Erk inhibitor, U0126; the Src inhibitor, PP2; and the Ras inhibitor, FTS, eliminated signaling in their specific pathway. However, PP2 also reduced Ras expression; and FTS slightly reduced Src phosphorylation. In addition, the VEGF-R2 inhibitor, SU5416, blunted Src activation; and the VDCC inhibitor, nifedipine, reduced Src phosphorylation and Ras expression. B, summary of expression levels and phosphorylation status of the signaling molecules in the presence of the pathway-specific inhibitors. Data are expressed as mean ± S.E. (n = 3 per condition).

Sublytic MAC Activation Increases VEGF Expression via the mRNA-destabilizing Protein Tristetraprolin (zfp36)

VEGF expression is tightly regulated. Hypoxia, in part through the transcription factor Hif1α, is thought to be the principal stimulus for VEGF secretion from the RPE (31); and immortalized human RPE cells have been shown to secrete VEGF in response to oxidative stress, with higher secretion toward the retina side (32). A second level of control is at the level of cytoplasmic stability of the mRNA encoding for VEGF. Stability of many cytokines, including VEGF, is controlled by AU-rich elements (ARE), a family of RNA sequences located within the 3′-untranslated region (3′-UTR) (33, 34). Specific binding proteins that bind to the ARE accelerate or slow down the decay of the respective transcript (35). VEGF mRNA has been shown to be destabilized by tristetraprolin or zfp36; and loss of zfp36 can result in increased VEGF stability and hence protein production (36). Here, we asked whether sublytic MAC activation affects VEGF and zfp36 mRNA levels, and whether those levels are affected by the intracellular signaling pathways activated by H₂O₂ + NHS. VEGF and zfp36 mRNA levels were measured over the first 4 h of exposure of RPE monolayers to H₂O₂ + NHS, or either H₂O₂ or NHS alone (Fig. 4A). Zfp36 mRNA levels increased rapidly after exposure to H₂O₂ + NHS, while VEGF levels were unaffected. During the 4 h of exposure time an inverse correlation was identified between zfp36 and VEGF. When the levels of the destabilizing protein zfp36 dropped, levels of VEGF mRNA increased. Exposure of monolayers with H₂O₂ or NHS alone for 4 h had no effect on the balance between zfp36 and VEGF mRNA levels; zfp36 mRNA levels remained elevated, while VEGF levels remained at baseline. As predicted based on the VEGF secretion data, cells exposed to H₂O₂ + NHS that were pretreated with Erk, Src, Ras, VEGF-R2, and VDCC inhibitors showed little increase in VEGF mRNA expression. While the levels of zfp36 mRNA levels were reduced in the inhibitor-treated when compared with cells exposed to H₂O₂ + NHS, levels were increased significantly by ∼4-fold when compared with untreated control cells (Fig. 4B). Collectively, these data suggest that zfp36 expression is associated with a reduction in VEGF expression, which in turn can alter the pathological phenotype of the RPE. However, additional experiments investigating zfp36 protein levels, phosphorylation status and mRNA and protein stability are needed to further characterize this interaction.

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FIGURE 4.

Complement activation on ARPE-19 cells induces the expression of VEGF mRNA and reduces expression of zfp36 mRNA, a known destabilizer of VEGF transcripts. Serum-deprived ARPE-19 cell monolayers with stable TER were treated with 0.5 mm H₂O₂ and 25% NHS in the presence and absence of pathway-specific inhibitors, and cells were collected after 4 h, or at the time points specified. VEGF and zfp mRNA levels were determined by QRT-PCR. A, exposure of monolayers to oxidative stress or serum resulted in an increase in zfp36, without affecting VEGF mRNA levels (3 h of exposure. Complement activation (H₂O₂ and NHS) however, led to a rapid increase in VEGF mRNA expression, and a concomitant decrease in zfp. B, pretreatment of cells exposed to H₂O₂ + NHS with Erk, Src, Ras, VEGF-R2, and VDCC inhibitors significantly blunted the increase in VEGF mRNA expression, and concomitantly increased zfp36 mRNA levels. On the other hand, cells pretreated with the P38 inhibitor had VEGF and zfp36 expression levels comparable to the H₂O₂ + NHS-treated controls. Data are expressed as mean ± S.E. (n = 3 per condition).

DISCUSSION

Our results demonstrate that sublytic MAC utilizes the regulated VEGF secretory pathway to increase VEGF release. Sublytic MAC was found to stimulate intracellular signaling molecules known to be part of the regulated secretory pathway such as Ras, Erk and Src, but not the basal VEGF secretory pathway, which is mediated by activation of P38 MAP kinase. Activity in the Erk/Ras and Src/VDCC pathways led to an increase in VEGF secretion followed by an impairment of transepithelial resistance; effects that could be prevented or significantly blunted by their respective inhibitors. Finally, activity in these pathways was correlated with a decrease in VEGF mRNA expression and a concomitant increase in the VEGF mRNA destabilizing protein, zfp36.

The complement system is an essential part of the evolutionarily ancient innate immune system, involved in eliminating foreign antigens and pathogens (reviewed in Ref. 37, 38). However, inappropriate or excessive complement activation is also involved in the pathogenesis of autoimmune, inflammatory, and ischemic disease states (reviewed in Ref. 39). The complement system can be activated by three distinct pathways: the classical (CP), lectin (LP), and alternative pathway (AP) (40). All three pathways lead to the generation of a C3 convertase enzyme complex. The convertases cleave C3 to produce C3a and a C3b fragment, and participate in the formation of C5 convertase, which cleaves C5 to yield C5b and the soluble anaphylatoxin, C5a. Formation of C5b initiates the terminal complement pathway resulting in the sequential assembly of complement proteins, C6, C7, C8, and C9, to form the cytolytic membrane attack complex (MAC or C5b-9). Triggering of this final pathway can lead to either lytic or sublytic activation. Lytic activation results in the assembly of the membrane attack complex, which is the cytolytic end-product of the complement cascade; it forms a transmembrane channel, causing osmotic lysis of the target cell. Sublytic activation occurs when the number of channels assembled on the cell surface is limited, which does not lead to the destruction of the target cell. At sublytic doses, the complement MAC complex has a wide range of effects on many cell types leading to changes in cellular responses such as secretion, adherence, aggregation, chemotaxis, cell division, or membrane function (reviewed in Ref. 41; examples in Refs. 42–45). Sublytic complement also induces increased cell resistance to lytic doses of complement. The transmembrane channel consists of the tetramolecular C5b-8 and tubular poly-C9 complex. The diameter of the pore created varies according to the amount of C9 molecules available to complex with bound C5b-8. Binding of the first molecule of C9 initiates C9 oligomerization at the site of MAC assembly; and once at least 12 molecules are incorporated into the complex, a discrete channel structure is formed (46). Human serum contains 42–77 μg/ml C9, which is 10–20-fold more than what is required for sublytic MAC activation; Sala-Sala-Newby et al. (47) have shown in a cell-based system using HeLa cells, that the threshold for sublytic MAC activation lies at ∼4 μg/ml of C9. The pore allows free movement of molecules in and out of the cell, since it has a hydrophilic internal face that allows the passage of water. Important for this work, Ca²⁺ influx has been shown to be one of the general consequences of MAC activation (48), resulting in Ca²⁺ oscillations that have been shown to last up to 45 min, in oligodendrocytes (49). In muscle cells, Jackson et al. (50) have shown using patch-clamp analysis of individual MAC channels, that these pores rapidly change between conducting and non-conducting states. Sublytic MAC activation is transient, exhibiting a half-life of ∼40 min, with MAC being removed from the cell surface by either outward or inward vesiculation (51). This process is Ca²⁺-calmodulin dependent (52). Finally, there are a number of different inhibitors that prevent MAC assembly. The membrane-bound, GPI-anchored inhibitor, CD59, acts by binding to the C8 and the C9 complement proteins, and thereby prevents formation of the lytic pore (53). In addition, the soluble inhibitors, S-protein (vitronectin) (54) or clusterin (complement-associated protein SP-40,40; complement cytolysis inhibitor; aging-associated gene 4 protein; Ku70-binding protein 1, NA1/NA2; testosterone-repressed prostate message 2; or apolipoprotein J) (55), which bind to the C5b-7 structure, prevent its attachment to cell membranes by rendering it water-soluble and lytic inactive. Thus, in ARPE-19 cells, due to the presence of complement inhibitors CD46, 55, and 59 on the cell surface, complement activation leads to transient MAC activation (18). The transient insertion of these transmembrane channels is expected to lead to Ca²⁺ influx, resulting in membrane depolarization and subsequent changes in VEGF secretion. Unfortunately, the calcium requirement could not be tested directly. Although the alternative pathway is active in the presence of Mg²⁺ EGTA, concentrations sufficient to eliminate Ca²⁺ in media containing 25% serum, significantly impaired TER. We expect to answer this question in future studies using calcium imaging, as well as patch clamp recordings to examine membrane potential and ion channel activity.

Overall, the data suggest the following possible mechanisms of VEGF secretion (Fig. 5). VEGF is typically present in granular vesicles (56) and exhibits polarized secretion in RPE cells (e.g. Ref. 29, 32) via a saturable process (57). Regulated secretion of polypeptides present in granular vesicles occurs via exocytosis triggered by Ca²⁺. Sublytic complement activation, which is due to the transient insertion of MAC, generating pores that allow the influx of ions, in particular Ca²⁺ and Na⁺, depolarizes the cell membrane. Membrane depolarization and the local influx of Ca²⁺ could therefore trigger the following consecutive or parallel events to result in the observed VEGF secretion. First, membrane depolarization is known to activate voltage-dependent calcium channels (VDCC), resulting in the local influx of Ca²⁺ required for VEGF release. This mechanism can be amplified by the MAC-mediated influx of Ca²⁺ leading to Src activation, which activates the VDCC. Interestingly, RPE cells from AMD patients show increased VEGF release via this Src/VDCC pathway (21). Second, Ca²⁺ influx is also known to activate the Ras/Erk pathway (58–60), a pathway that has been shown to be involved in the regulated secretion of VEGF (22). Because Erk activity has been shown to result in a transient rise in intracellular Ca²⁺ (61), this might be the source of Ca²⁺ required to secrete VEGF via the Ras/Erk pathway. Thus, there seems to be at least two intracellular signaling pathways that are triggered by sublytic MAC formation, Src/VDCC and Ras/Erk. Based on the transient nature of the increase in Erk phosphorylation and Ras expression levels, and the sustained increase in Src phosphorylation, the two mechanisms together might be responsible for an initial burst followed by a more sustained level of VEGF secretion. Activity in these two pathways might mediate the VEGF-induced reduction in TER, which plateaus within 1–4 h after stimulation (18, 29). The inhibitor experiments presented in Fig. 3 suggest that there is crosstalk between these two pathways at the level of Src and Ras. Src-mediated signaling via Ras has been reported in other systems (62, 63); however, signaling in the other direction appears to be less common. In summary, sublytic MAC-mediated VEGF secretion occurs via two known pathways that control regulated, rather than basal VEGF release.

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FIGURE 5.

Sublytic MAC activation in RPE-cells increases regulated VEGF secretion. VEGF, which is present in granular vesicles, is secreted via exocytosis triggered by Ca²⁺. Sublytic complement activation, results in the influx of Ca²⁺ and Na⁺ ions, which triggers both Ca²⁺-activated pathways and also depolarizes the cell membrane. Membrane depolarization activates voltage-dependent calcium channels (VDCC), allowing for the local influx of Ca²⁺ required for exocytosis. VDCC activation can be amplified by MAC-mediated influx of Ca²⁺ leading to Src activation. Ca²⁺ influx also activates the Ras/Erk pathway known to be involved in the regulated secretion of VEGF. Thus, sublytic MAC formation engages at least two intracellular signaling pathways that result in regulated VEGF secretion: Src/VDCC and Ras/Erk. The inhibitor experiments also revealed crosstalk between these two pathways. Finally, both Ras- and Erk-mediated signaling have been reported to inhibit zfp, which in turn results in increased VEGF expression. Thus, we hypothesize that sublytic MAC-activation engages pathways involved in regulated VEGF secretion rather than basal VEGF release.

As outlined above, the expression of angiogenic factors such as VEGF is carefully regulated, involving the tightly balanced transcription factor, Hif1α (31), and a mechanism that controls its mRNA stability through the mRNA destabilizing protein, zfp36 (36). Here, we reported on two observations. First, exposing serum-deprived RPE monolayers (serum was withdrawn in all experiments 5–7 days prior to the experiment to allow for complement activation with 25% serum) to H₂O₂, NHS, or H₂O₂ + NHS resulted in a rapid increase in zfp36 without affecting VEGF mRNA levels. Zfp36 has been shown to be induced by a variety of conditions, including growth factor present in serum (64) and oxidative stress (65). Second, we showed that sublytic MAC-activation had opposing effects on VEGF and zfp36 mRNA levels; H₂O₂ + NHS stimulation increased VEGF and decreased zfp36 mRNA levels, whereas the inhibitors to those pathways involved in stimulating VEGF secretion, had the opposite effects. It has been reported that Erk activity is associated with a decrease in zfp36 activity (36, 66), linking Erk activity with VEGF mRNA expression. In addition, it has been shown that both Src and Ras (66) can activate VEGF expression, presumably through the phosphorylation of nuclear transcription factors; and since phosphorylated zfp36 is less active, this may be an alternative means to regulate VEGF levels in the MAC-stimulated RPE cells. However, additional experiments investigating zfp36 protein levels, phosphorylation status, and mRNA and protein stability are needed to further distinguish between these possibilities.

VEGF is an important signaling protein involved in both de novo formation of the embryonic circulatory system and angiogenesis, and signals via two tyrosine kinase transmembrane VEGF receptors, flt-1 and flk-1 (67). The RPE is a major source of VEGF (e.g. Ref. 68). Histopathological studies of CNV membranes from patients with AMD have demonstrated the presence of VEGF and its receptors (e.g. Ref. 69). Animal studies support a role for increased VEGF of RPE origin in CNV progression (70). Given the importance of VEGF, currently approved therapies are directed at CNV, which occurs in patients with wet AMD. Current treatments (Macugen and Lucentis), directed at inhibiting VEGF, have been shown to improve vision and/or slow progression of the disease (71). However, since basal levels of VEGF may be required for maintenance of the adult choroidal vasculature, and VEGF is a critical survival factor in the retina (72), long-term anti-VEGF therapy may not be safe. Thus, focusing on a treatment that reduces the pathologic, rather than the basal VEGF expression may be essential for maintaining normal blood vessel health. Hence, therapeutic approaches targeting the regulated VEGF secretory pathway to reduce VEGF release and production should be investigated; and since the alternative pathway of complement controls the regulated VEGF secretory pathway, interfering at the level of complement to reduce inflammation and pathological VEGF secretion (e.g. Ref. 73) might provide a unique strategy for the treatment of AMD.

Acknowledgments

We thank Joshua Thurman (University of Colorado Denver School of Medicine, Denver, CO) for helpful discussions and Luanna Bartholomew for critical review (Medical University of South Carolina, Charleston, SC). Animal studies were conducted in a facility constructed with support from the National Institutes of Health (C06 RR015455).

^*This work was supported, in whole or in part, by Grant R01 EY019320 from the National Institutes of Health, Grant I01 RX000444 from the Dept. of Veterans Affairs, Foundation Fighting Blindness, and an unrestricted grant to Medical University of South Carolina from Research to Prevent Blindness, Inc.

²The abbreviations used are:

AMD

age-related macular degeneration

RPE

retinal pigment epithelium

TER

trans-epithelial resistance

VEGF

vascular endothelial growth factor

MAC

membrane attack complex

VDCC

voltage-dependent calcium channel

CSS

complement-sufficient serum

CFB

complement factor B

CFH

complement factor H

C1INH

complement 1 inhibitor

CNV

choroidal neovascularization.

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AtomRemote

Wireless Remote Control

The AtomRemote app for iOS and macOS provides external controls for NINJA V, and NINJA V+ with AtomX SYNC module. This app enables filmmakers to perform a range of configuration tasks and operations wirelessly up to 15 meters via Bluetooth LE. Input options include the ability to define how the device connects to the camera, select Gamma/EOTF and adjust Gamut settings. For monitoring, AtomRemote can be used to control playback, select monitoring modes, apply custom 3D LUTs and analysis tools including exposure and focus. Output controls provide options for 4K to HD and LUT preview and HDR output.

Learn More About AtomX SYNC

Instant Playback & Review

Ninja V takes exactly what your camera sensor is capturing, enhances it and allows you to view and record it in amazing detail. Ninja V gives you the ability to record and instantly playback, review and tag your favourites takes to create playlists. This portable powerhouse gives you complete confidence in what you’re capturing, so you’re free to push creative boundaries more than ever before.

 

ProRes RAW on
Canon EOS R5 C and Ninja V

The Ninja V supports recording of ProRes RAW up to 6K at up to 30 fps and 3K at up to 60fps from the Canon R5 C mirrorless camera. The EOS R5 C features a new heat dissipation system allows for extended takes and the most demanding production days.

The efficiencies of ProRes RAW make it possible to playback multiple streams of 8Kp30 ProRes RAW simultaneously. This makes it perfect for multicam projects. The built-in LTC input on the EOS R5 C means that it’s also suited for multicamera productions. It can be used with the Atomos UltraSync One, a compact wireless sync device, which can be used to connect and synchronise multiple cameras.

The technology is also compatible with the AtomX SYNC module for the Ninja V/V+. This means you can wirelessly synchronise the EOS R5 C to a range of devices. Coupled with the established Canon EOS lens systems and powerful Servo AF technology, this combination will propel filmmakers to a whole new level in terms of image quality and production value.

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RECORD RAW

Ninja V & ProRes RAW

The number of cameras which now support RAW recording over HDMI on the Ninja V continues to grow. ProRes RAW combines the visual and workflow benefits of RAW video with the incredible real-time performance of ProRes. This format gives filmmakers enormous latitude when adjusting the look of their images, making it ideal for HDR workflows.

COMPATIBLE CAMERAS

ProRes RAW Recording with Ninja V

Learn more about Ninja V and the latest cameras at Atomos Academy

Optional accessories for your Ninja V

Essential accessories for capture, post-production and streaming

• AtomX SDI

  Input 12G/6G/3G/HD with the ability to toggle between input channel sources on SDI 1 / 2. Supporting up to 4kp60. Dual-link support for 1.5G/3G and 6G.

• Battery Eliminator

  Power select Atomos monitor/recorder units

• Sunhood

  Block excess reflections from washing out the displayed image

• SSD

  Custom made for the new Atomos Ninja V monitor recorder

• Docking Station

  Offload the drive contents direct to your computer

• Connect

  Bridge recording and monitoring with streaming platforms such as YouTube, Twitch and OBS.

• Monitor Mount

  Features a quick release plate, as well as 360° pan and 180° tilt functionality

• DTap Cable

  D-Tap to DC Locked connector Barrel Coiled Cable for Battery Eliminator

• AtomX Arm

  Attach your Atomos monitor to your camera cage or rig via a quick release workflow.

PRO MONITORING

Access All Areas

The Ninja V offers a huge range of features that can enhance your workflow, from AtomOS Monitor Assist features like Focus Peaking, Zoom, Vectorscopes &  False Color through to Safe Areas, Cine Guides and our AtomHDR processing engine that allows realtime Log>PQ/HLG previewing and more. Customizable settings & view modes — and instantly clear all overlays with a simple touch of the screen. Take control of your production at capture with Atomos Ninja V.

Focus Peaking
Customizable

Weddings. Parties. Anything.

Universally popular, Ninja V ticks more boxes for more filmmakers than any other 5” monitor on the market. And it will only get better. The Ninja V’s features continue to improve and evolve with every new AtomOS release. Not only will additional ProRes RAW capable cameras be supported, other new features are constantly being added for free.

Recent additions include a wider range of frame guides (1:1, 4:5, 1:91:1 and 9:16) – perfect for social media content creation. A nine-grid overlay makes composing using the rule of thirds easy. The addition of an ARRI style false colour scale bolsters Ninja V’s impressive professional credentials.

Lock the shot with Ninja V’s monitoring and composition essentials.

• Social Guides

  1:1 / 4:5 / 1:91:1 / 9:16

• Safe Areas

  TV / Screen Guides

• Frame Guides

  SMPTE standards

Learn, Inspire, Create.

 

Check out the latest content created using Atomos Ninja V.
Features from Creators, tips from Pros and the latest innovations from Atomos.

BROWSE ACADEMY

SSD MEDIA

Break through the recording time barrier

Shooting 4K and above in advanced formats like ProRes, DNx and ProRes RAW requires storage media with ample volume, fast transfer times and high, sustained read and write speeds.

AtomX SSDmini drives complete the Ninja V’s digital workflow. Smaller than conventional SATA SSDs they are an affordable professional alternative to recording to a camera’s internal memory cards. Offering up to 2TB storage, a sequential read speed of up to 550MB/s and write speed up to 500MB/s, these drives can record up to 150 minutes of 4K ProRes on a single drive. Measuring 8cm long, 7.5cm wide and weighing as little as 88g, the custom built drives neatly fit the Ninja V’s compact proportions.

Fast. Reliable. Compact. Rugged.

ProRes RAW

Olympus E-M1X & E-M1 MIII with the Ninja V

WATCH: Janne Amunét Director from Kauas Creative speaks first-hand of his experience shooting in ProRes RAW on the Olympus E-M1X & E-M1 MIII with the Ninja V. Understand the key features that make up the Olympus E-M1X & E-M1 MIII such as weatherproofing and image stabilisation and what this means when the mirrorless cameras are paired with the Ninja V to shoot in ProRes RAW.

 

Related Posts

FEATURE

ProRes RAW on Z CAM + Ninja V

WATCH: Go behind the scenes of UN/SEEN, an artistic abstract short film by filmmaker/director James Tonkin. UN/SEEN was shot on the Z CAM E2-F6 full-frame cinema camera and the Atomos Ninja V HDR monitor-recorder in Apple ProRes RAW.

The behind the scenes video was shot on the Z CAM E2 and Ninja V in ProRes RAW. Watch the finished video below. Then view final project inHDR.

 

ATOMOS ACADEMY

ATOMX SYNC

Making multi-camera mainstream

You no longer need a huge budget or a highly skilled editor to produce perfectly synchronised, multicamera video. The optional AtomX SYNC module sits on the back of the Ninja V, integrating it into a Timecode Systems powered multi-camera sync network. The advanced long-range RF system is completely wireless, offering incredible accuracy and stability. Effortlessly synchronise your recordings with multiple camera and sound sources. With frame-accurate timecode embedded directly into each recording you can accurately edit together multiple video and audio sources effortlessly using automated functions in all popular editing software.

LEARN MORE

CALIBRATION

Trust what you see

Ninja V is your go anywhere monitoring system. Any camera, anywhere, any time of day, you can trust that you’re accurately seeing what you are recording. To ensure reliable and dependable accuracy, Atomos have partnered with calibration leader X-Rite to ensure all of our monitors operate seamlessly across Log/HDR capture, Post Production and HRD/SDR delivery. Ninja V is easy to calibrate, ensuring you are seeing your images accurately.

 

X-RITE i1

Unlock the full potential of your camera

The diversity of codecs makes the Ninja V compatible with all major editing software packages. In terms of capture, virtually any HDMI or SDI source is supported, including cameras from Nikon, FujiFilm, Canon, Panasonic, Sony, Z-Cam, Olympus, RED and ARRI.

Class leading monitoring technology

• 60-240hz

  Recording frequency

• Auto HDR

  Flags for TV setup

• Zoom

• Focus Peaking

• LUTS

• Zebra

Gaming

Record 4K HDR Gaming

Ninja V is a stand-alone system for 4k UHD, HDR and high frame rate capture that eliminates expensive, complicated and unreliable PC setups. Ninja V enables simple recording, monitoring and instant review. Capture every detail in HDR and automatically include all the correct HDR flags ready for upload to YouTube. Ideal for games development testing, pre-release capture sessions or just to show off your skills!

Ninja V

Universally popular, Ninja V ticks more boxes for more filmmakers than any other 5” monitor on the market.

$

Please note:
Specifications are subject to change without notice.
All information correct at time of publishing.
Price above is for a limited time only.

° Paid activation. Available at myatomos.com

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