Name: using an automated hirschberg test app to evaluate ocular alignment
Uploaded: 2020-03-24T15:00:00
Description: Watch this Scientific Journal Video about Using an Automated Hirschberg Test App to Evaluate Ocular Alignment at JoVE.com

Dette arbeidet ble delvis støttet av NIH-stipendet R44EY025902 og av Mass Eye & Ear Curing Kids Grant.

Denne studien ble utført i samsvar med tenets av erklæringen av Helsinki, ved Schepens Eye Research Institute (Boston, MA) og Spaulding Rehabilitation Hospital (Boston, MA). Informert samtykke ble innhentet fra alle deltakerne. Studien ble godkjent av de lokale institusjonelle gjennomgangsrådene i Mass Eye and Ear (Boston, MA).
MERK: Kriterier for pasientinklusjon var tidligere diagnose av horisontal esotropi (konstant eller intermitterende exotropi eller esotropi) og ingen andre synshemmin...

<ol>
	<li>Kemper, A. R., Clark, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preschool+vision+screening+in+pediatric+practices.">Preschool vision screening in pediatric practices.</a> Clinical Pediatrics. 45 (3), 263-266 (2006).</li><li>Rowe, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+profile+of+strabismus+in+stroke+survivors.">The profile of strabismus in stroke survivors.</a> Eye. 24 (4), 682-685 (2010).</li><li>Black, K., McCarus, C., Collins, M. L. 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A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fixation+preference+and+visual+acuity+testing+in+a+population-based+cohort+of+preschool+children+with+amblyopia+risk+factors.">Fixation preference and visual acuity testing in a population-based cohort of preschool children with amblyopia risk factors.</a> Ophthalmology. 116 (1), 145-153 (2009).</li><li>Robaei, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+Associated+with+Childhood+Strabismus:+Findings+from+a+Population-Based+Study.">Factors Associated with Childhood Strabismus: Findings from a Population-Based Study.</a> Ophthalmology. 113 (7), 1146-1153 (2006).</li><li>Tarczy-Hornoch, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Risk+factors+for+decreased+visual+acuity+in+preschool+children:+the+multi-ethnic+pediatric+eye+disease+and+Baltimore+pediatric+eye+disease+studies.">Risk factors for decreased visual acuity in preschool children: the multi-ethnic pediatric eye disease and Baltimore pediatric eye disease studies.</a> Ophthalmology. 118 (11), 2262-2273 (2011).</li><li>Chan, K. W., Deng, L., Weissberg, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+of+Strabismus+by+Non-Health+Care+Professionals+in+an+Ethnically+Diverse+Set+of+Images.">Detection of Strabismus by Non-Health Care Professionals in an Ethnically Diverse Set of Images.</a> JAMA Ophthalmology. 134 (1), 30-36 (2016).</li><li>Hunter, D. G., Guyton, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vertical+location+of+the+corneal+light+reflex+in+strabismus+photography.">Vertical location of the corneal light reflex in strabismus photography.</a> Archives of Ophthalmology. 116 (6), 767-771 (1998).</li><li>Eskridge, J. B., Wick, B., Perrigin, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Hirschberg+test:+a+double-masked+clinical+evaluation.">The Hirschberg test: a double-masked clinical evaluation.</a> American Journal of Optometry and Physiological Optics. 65 (9), 745-750 (1988).</li><li>Hasebe, S., Ohtsuki, H., Tadokoro, Y., Okano, M., Furuse, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+reliability+of+a+video-enhanced+Hirschberg+test+under+clinical+conditions.">The reliability of a video-enhanced Hirschberg test under clinical conditions.</a> Investigative Ophthalmology &amp; Visual Science. 36 (13), 2678-2685 (1995).</li><li>Pundlik, S., Tomasi, M., Liu, R., Houston, K., Luo, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+and+Preliminary+Evaluation+of+a+Smartphone+App+for+Measuring+Eye+Alignment.">Development and Preliminary Evaluation of a Smartphone App for Measuring Eye Alignment.</a> Translational Vision Science &amp; Technology. 8 (1), 19 (2019).</li><li>Cheng, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+EyeTurn+App+for+School+Vision+Screening.">The EyeTurn App for School Vision Screening.</a> American Academy of Optometry Annual Meeting. , (2018).</li><li>Committee on Practice and Ambulatory Medicine and Section on Ophthalmology. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+Photoscreening+for+Children's+Vision+Screening.">Use of Photoscreening for Children's Vision Screening.</a> Pediatrics. 109 (3), 524-525 (2002).</li><li>Simons, B. D., Siatkowski, R. M., Schiffman, J. C., Berry, B. E., Flynn, J. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pediatric+photoscreening+for+strabismus+and+refractive+errors+in+a+high-risk+population.">Pediatric photoscreening for strabismus and refractive errors in a high-risk population.</a> Ophthalmology. 106 (6), 1073-1080 (1999).</li><li>Kerr, N. C., Somes, G., Enzenauer, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+developmentally-at-risk+status+on+the+reliability+of+the+iScreen(R)+photorefractive+device+in+young+children.">The effect of developmentally-at-risk status on the reliability of the iScreen(R) photorefractive device in young children.</a> The American Journal of Orthopedics. 61, 117-123 (2011).</li><li>Arnold, R. W., O'Neil, J. W., Cooper, K. L., Silbert, D. 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En person uten profesjonell opplæring kan bruke EyeTurn-appen til å ta bilder av øynene og oppnå okulære justeringsmålinger, som kan tolkes av en øyespesialist på stedet eller eksternt. Appen gir bare omfanget av feiljusteringen, i stedet for noen tolkning eller diagnose. Øyepleiefagfolk som øyeleger eller øyeleger bør avgjøre om feiljusteringen er betydelig eller ikke, og foreta en diagnose etter å ha vurdert andre faktorer, inkludert forholdene som målingen ble tatt under.
Å ...

Måling av øyejustering utføres ofte i visjonsklinikker. Dekktest med prismenøytralisering er den vanlige kliniske metoden for å kvantifisere graden av øyefeiljustering (strabismus). Denne metoden krever en høy grad av trening og erfaring. Nøyaktig måling blir mer utfordrende når pasienter ikke fullt ut kan engasjere seg i eksamen som små barn1, personer med hjerneskader eller hjerneslag2, eller utviklingshemming3. Videre er det behov for okulær justeringstesting i skolescreening, fordi strabismus utvikler seg i barndommen i anslagsvis 5-8% av den amerikanske befolkningen4, og er en betydelig risikofaktor for amblyopi med ca 30-40% av tilfellene av amblyopi tilskrives strabismus5,6,7. Skolesykepleiere er imidlertid normalt ikke opplært til å gjennomføre standard dekktest med prismenøytralisering for slik screening. For ikke-øye-omsorg fagfolk, en ekstra utfordring i strabismus screening er at intermitterende strabismus (feiljustering er ikke alltid manifestert) og mindre størrelser av feiljustering er ikke visuelt åpenbare (<15 prisme diopters [Δ])8.
I et forsøk på å løse utfordringene ved deteksjon og måling av skjeling, har vi utviklet en smarttelefonapp (EyeTurn) som implementerer og automatiserer den fotografiske Hirschberg-metoden9 ved å sammenligne forskyvningen av hornhinnenrefleksjoner mellom øynene. Mens konvensjonell fotografisk Hirschberg metoden har vist seg å ha god reproduserbarhet i klinikker10,11, kostnaden for dedikerte, frittstående enheter er en barriere for bred adopsjon. Ved å gi et brukervennlig verktøy for å måle øyejustering med standard smarttelefoner, hypoteser vi det vil bli mye vedtatt i skolen visjon screening og brukes av ikke-øyeomsorg fagfolk. Våre tidligere evalueringsstudier har vist at appmålingen er i samsvar med gjeldende klinisk standard for prisme og alternativ dekktest12, for strabismus størrelser av esotropi og eksotropi opptil 60Δ. I en pilotskolescreeningsstudie viste vi også at appen kan hjelpe skolesykepleieren med å oppdage barn med intermitterende exotropi som ble savnet av standard skolevisjonsscreeningprotokoller13.
IOS-versjonen av appen er for tiden tilgjengelig for forskere og klinikere på forespørsel om forskningsformål. Bestillerne har så langt inkludert skolesykepleiere, pediatriske øyeleger, øyeleger, nevro-øyeleger og strabismus spesialister. Formålet med denne artikkelen er å dele de detaljerte appprotokollene for bruk av appen for å evaluere okulær justering under forskjellige visningsforhold, nemlig nær og fjern fikseringsavstand; med og uten øyebelegg for å bryte kikkertfusjon.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>EyeTurn</td>
			<td>EyeNexo</td>
			<td></td>
			<td>Smartphone app for measureing eye misalignment</td>
		</tr>
	</tbody>
</table>

using an automated hirschberg test app to evaluate ocular alignment

En smarttelefonapp er utviklet for å utføre den automatiserte fotografiske Hirschberg-testen for objektiv måling av okulær feiljustering. Ved å beregne forskjellen i hornhinnerefleksjon generert av telefonkameraet blinker i forhold til irissenteret basert på høyoppløselige bilder, kan appen måle feiljustering med en mye høyere presisjon enn det blotte øye som utfører Hirschberg-testen. Det har blitt validert i en tidligere klinisk evalueringsstudie ved å sammenligne med den kliniske gullstandardprisme og alternativ dekktest. Målet med denne artikkelen er å beskrive testteknikkene for hvordan du bruker appen til å måle okulær justering for ulike fikseringsavstander, uten eller med deksel for å bryte fusjon, samt vinkelkappa, slik at brukerne kan bruke appen til å utføre tilsvarende tester som vanligvis gjøres på klinikken ved hjelp av prismer.

I dette arbeidet beskriver vi protokollene for å evaluere okulær justering ved hjelp av en smarttelefonapp som utfører den fotografiske Hirschberg-testen. Grensesnittet til appen vises i figur 1. Brukerne kan velge å utføre covertest eller måle en pasient med begge øynene fiksering på et mål samtidig, enten på nær eller langt fiksering avstander. Når visningsforholdene er bestemt avhengig av testformålene, kan brukerne følge protokollene og ta e...

Watch this Scientific Journal Video about Using an Automated Hirschberg Test App to Evaluate Ocular Alignment at JoVE.com

Using an Automated Hirschberg Test App to Evaluate Ocular Alignment

Vi presenterer en protokoll for å bruke en smarttelefonapp til å utføre Hirschberg-test for måling av manifest og intermitterende okulær feiljustering (strabismus) under nær og langt fikseringsforhold.

Bruke en automatisert Hirschberg-testapp til å evaluere okulær justering

A smartphone app has been developed to perform the automated photographic Hirschberg test for objective measurement of ocular misalignment. By computing ...

Mobilappen vi har utviklet kan brukes til å måle skjeling for forskning eller til klinisk bruk. Denne metoden forenkler en enkel skjelingsmåling, som for tiden krever omfattende opplæring og erfaring som skal utføres ved hjelp av bare et standard telefonkamera og appprogramvaren. I appgrensesnittet kan brukerne velge å utføre en dekktest eller måle en pasient med begge øynene fiksert på et mål samtidig, enten på en nær eller langt fikseringsavstand.For å måle tropia, med ett enkelt øyeblikksbilde i med en nær fiksering, start appen og sett modusen til No Cover. Velg I nærheten og plasser telefonen i liggende retning ca. 40 centimeter fra pasienten i øyenivå, med det bakre kameraet vendt mot pasienten. Be pasienten om å fiksere på lommelykten, som skal være av på dette punktet.For tester som krever en nøyaktig innkvartering, lim inn et fikseringsmål på baksiden av telefonen rett under eller over lommelykten. Når pasienten bekrefter at de fikserer, trykker du på den runde knappen for å ta et øyeblikksbilde som skal analyseres av appen. Når analysen er fullført, vil appen vise den oppdagede limbusgrensen, øyets sentrum og plasseringen av hornhinnens refleksjon.Etter å ha bekreftet at disse funksjonene har blitt oppdaget uten åpenbare feil, observere målingene knyttet til øyejusteringen. Trykk deretter på Lagre-knappen for å lagre den gjeldende testen i telefonen, eller trykk på pil tilbake-knappen for å teste på nytt. Hvis du vil måle uregelmessig okulær feiljustering for fjernfiksering i appen, setter du modusen til Dekktest og velger Langt.Hold telefonen i en liggende retning, ca 40 centimeter fra pasienten i øyenivå med blitskameraet plassert mellom pasientens øyne. Be pasienten om å se like over telefonen, til et mål omtrent seks meter unna. Få pasienten til å dekke ett øye og trykke på den runde knappen, vil appen begynne deteksjonen.Mens du sørger for at pasienten fikserer riktig, må du få dem til å fjerne okklusjonen raskt. Appen tar automatisk et bilde så snart okklusjonen er fjernet. Når analysen er fullført, vil appen vise den oppdagede limbusgrensen, øyets sentrum og plasseringen av hornhinnens refleksjon.Etter å ha bekreftet at disse funksjonene oppdages uten åpenbare feil, gjennomgår du måleresultatene for øyejusteringen i prisme diopters, før du lagrer dataene eller gjentar testen. For Angle Kappa-måling i appen velger du mål Kappa-vinkel. Plasser telefonen i en liggende retning ca 40 centimeter fra pasienten på øyenivå, og be pasienten om å bruke øyet som skal testes for å fiksere på lommelykten mens den dekker det andre øyet.Mens du sørger for at pasienten fikserer riktig, trykker du på den runde knappen for å ta et øyeblikksbilde, som vil bli analysert av appen. Når analysen er fullført, vil appen vise den oppdagede limbusgrensen, øyets sentrum og plasseringen av hornhinnens refleksjon. Etter å ha bekreftet at disse funksjonene oppdages uten åpenbare feil, kan du se måleresultatene for Kappa vinkel i grader.I denne representative analysen ble limbusgrensene til de to øynene og hornhinnens refleksjon av lommelykten oppdaget riktig, noe som tyder på at okulær justeringsmål ikke er gjenstand for bildeanalysefeil. I dette spesielle tilfellet hadde pasienten forlatt eksotropi som det er åpenbart fra den større hornhinnen refleksjon offset i venstre øye. Appen rapporterte ikke hvilket øye som ble avveket, fordi i tilfeller av en liten skjelingvinkel og ukjent Kappa-vinkel, ville det være upålitelig for appen å bestemme det avvikede øyet.Til sammenligning vises et eksempel uten skjeling. I denne analysen kan feilaktig limbusdeteksjon observeres. Mens påvisning av hornhinnen refleksjon er riktig, den grønne sirkelen klart ikke samsvarer med limbus grensen, noe som indikerer at testen skal gjentas.I denne analysen av 14 forsøkspersoner viste 10 personer eksotropi og fire forsøkspersoner presentert med eksotropi. Lineær regresjonsanalyse viste at øyemålingene av skjelingvinklene var konsistente med kliniske dekktestmålinger. Når du bruker appen, er det to viktige ting å huske på.En, sett dekktesten og fikseringsmodusen riktig for hver bestemt test. To, sjekk om øyefunksjonsdeteksjonen er riktig. Appen tillater bare skjelingsmåling, og det gir ikke en diagnose.Klinikere må kombinere okulær måling med andre synsmålinger for å gjøre en diagnose.

using-an-automated-hirschberg-test-app-to-evaluate-ocular-alignment

Research

JoVE Journal

Medicine

An in vivo Assay to Test Blood Vessel Permeability

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under physiologic conditions the endothelium is impermeable to albumin, so Evans blue bound albumin remains restricted within blood vessels. In pathologic conditions that promote increased vascular permeability endothelial cells partially lose their close contacts and the endothelium becomes permeable to small proteins such as albumin. This condition allows for extravasation of Evans Blue in tissues. A healthy endothelium prevents extravasation of the dye in the neighboring vascularized tissues. Organs with increased permeability will show significantly increased blue coloration compared to organs with intact endothelium. The level of vascular permeability can be assessed by simple visualization or by quantitative measurement of the dye incorporated per milligram of tissue of control versus experimental animal/tissue. Two powerful aspects of this assay are its simplicity and quantitative characteristics. Evans Blue dye can be extracted from tissues by incubating a specific amount of tissue in formamide. Evans Blue absorbance maximum is at 620 nm and absorbance minimum is at 740 nm. By using a standard curve for Evans Blue, optical density measurements can be converted into milligram dye captured per milligram of tissue. Statistical analysis should be used to assess significant differences in vascular permeability.

An in vivo Assay to Test Blood Vessel Permeability

We are presenting an in vivo assay to test blood vessel permeability. This assay is based on intravenous injection of a dye and subsequent visualization of its diffusion into interstitial spaces.

En<em&gt; In vivo</em&gt; Assay til Test Blood Vessel Permeabilitet

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under ...

Using Quantitative Real-time PCR to Determine Donor Cell Engraftment in a Competitive Murine Bone Marrow Transplantation Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules that regulate HSCs. In these transplant model systems, the function of HSCs is determined by the ability of these cells to engraft and reconstitute lethally irradiated recipient mice. Commonly, the donor cell contribution/engraftment is measured by antibodies to donor- specific cell surface proteins using flow cytometry. However, this method heavily depends on the specificity and the ability of the cell surface marker to differentiate donor-derived cells from recipient-originated cells, which may not be available for all mouse strains. Considering the various backgrounds of genetically modified mouse strains in the market, this cell surface/ flow cytometry-based method has significant limitations especially in mouse strains that lack well-defined surface markers to separate donor cells from congenic recipient cells. Here, we reported a PCR-based technique to determine donor cell engraftment/contribution in transplant recipient mice. We transplanted male donor bone marrow HSCs to lethally irradiated congenic female mice. Peripheral blood samples were collected at different time points post transplantation. Bone marrow samples were obtained at the end of the experiments. Genomic DNA was isolated and the Y chromosome specific gene, Zfy1, was amplified using quantitative Real time PCR. The engraftment of male donor-derived cells in the female recipient mice was calculated against standard curve with known percentage of male vs. female DNAs. Bcl2 was used as a reference gene to normalize the total DNA amount. Our data suggested that this approach reliably determines donor cell engraftment and provides a useful, yet simple method in measuring hematopoietic cell reconstitution in murine bone marrow transplantation models. Our method can be routinely performed in most laboratories because no costly equipment such as flow cytometry is required.

Determining donor cell engraftment presents a challenge in mouse bone marrow transplant models that lack well-defined phenotypical markers. We described a methodology to quantify male donor cell engraftment in female transplant recipient mice. This method can be used in all mouse strains for the study of HSC functions.

Bruke kvantitativ real-time PCR fastslår Donor Cell engraftment i en konkurransedyktig Murine benmargstransplantasjon Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules ...

Using a Knee Arthrometer to Evaluate Tissue-specific Contributions to Knee Flexion Contracture in the Rat

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ROM is called a joint contracture and results in increased morbidity. Due to the difficulty of reversing established knee contractures, early detection is important, and hence, knowing risk factors for their development is essential. The rat represents a good model with which the effect of an intervention can be studied due to the similarity of rat knee anatomy to that of humans, the rat's ability to tolerate long durations of knee immobilization in flexion, and because mechanical data can be correlated with histologic and biochemical analysis of knee tissue.
Using an automated arthrometer, we demonstrate a validated, precise, reproducible, user-independent method of measuring the extension ROM of the rat knee joint at specific torques. This arthrometer can be used to determine the effects of interventions on knee joint ROM in the rat.

The goal of the protocol is to measure the extension range of motion of the rat knee. The effects of various diseases that increase the stiffness of the knee joint and the effectiveness of treatments can be quantified.

Bruke en kne Arthrometer å vurdere vev-spesifikke bidrag til kneet strekking Contracture i rotte

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ...

Sucrose Preference and Novelty-Induced Hypophagia Tests in Rats using an Automated Food Intake Monitoring System

The prevalence and incidence of depressive disorders are rising worldwide, affecting about 322 million individuals, underlining the need for behavioral studies in animal models. In this protocol, to study depression-like and anhedonic behavior in rats, the established sucrose preference and novelty-induced hypophagia tests are combined with an automated food and liquid intake monitoring system. Prior to testing, in the sucrose preference paradigm, male rats are trained for at least 2 days to consume a sucrose solution in addition to tap water. During the test, rats are again exposed to water and sucrose solution. Consumption is registered every second by the automated system. The ratio of sucrose to total water intake (sucrose preference ratio) is a surrogate parameter for anhedonia. In the novelty-induced hypophagia test, male rats undergo a training period in which they are exposed to a palatable snack. During training, rodents show a stable baseline snack intake. On test day, the animals are transferred from home cages into a fresh, empty cage representing a novel unknown environment with access to the known palatable snack. The automated system records the total intake and its underlying microstructure (e.g., latency to approaching the snack), providing insight into anhedonic and anxious behaviors. The combination of these paradigms with an automated measuring system provides more detailed information, along with higher accuracy by reducing measuring errors. However, the tests use surrogate parameters and only depict depression and anhedonia in an indirect manner.

Presented here is a protocol to study depression-like and anhedonic behavior in rats. It combines two well-established behavioral methods, the sucrose preference and novelty-induced hypophagia tests, with an automated food and liquid intake monitoring system, to indirectly investigate rodent behavior using surrogate parameters.

Sukrose preferanse og nyhet-indusert hypofagi tester i rotter ved hjelp av en automatisert matinntak overvåkingssystem

The prevalence and incidence of depressive disorders are rising worldwide, affecting about 322 million individuals, underlining the need for behavioral ...

Endothelial Cell Transcytosis Assay as an In Vitro Model to Evaluate Inner Blood-Retinal Barrier Permeability

Dysfunction of the blood-retinal barrier (BRB) contributes to the pathophysiology of several vascular eye diseases, often resulting in retinal edema and subsequent vision loss. The inner blood-retinal barrier (iBRB) is mainly composed of retinal vascular endothelium with low permeability under physiological conditions. This feature of low permeability is tightly regulated and maintained by low rates of paracellular transport between adjacent retinal microvascular endothelial cells, as well as transcellular transport (transcytosis) through them. The assessment of retinal transcellular barrier permeability may provide fundamental insights into iBRB integrity in health and disease. In this study, we describe an endothelial cell (EC) transcytosis assay, as an in vitro model for evaluating iBRB permeability, using human retinal microvascular endothelial cells (HRMECs). This assay assesses the ability of HRMECs to transport transferrin and horseradish peroxidase (HRP) in receptor- and caveolae-mediated transcellular transport processes, respectively. Fully confluent HRMECs cultured on porous membrane were incubated with fluorescent-tagged transferrin (clathrin-dependent transcytosis) or HRP (caveolae-mediated transcytosis) to measure the levels of transferrin or HRP transferred to the bottom chamber, indicative of transcytosis levels across the EC monolayer. Wnt signaling, a known pathway regulating iBRB, was modulated to demonstrate the caveolae-mediated HRP-based transcytosis assay method. The EC transcytosis assay described here may provide a useful tool for investigating the molecular regulators of EC permeability and iBRB integrity in vascular pathologies and for screening drug delivery systems.

Endothelial Cell Transcytosis Assay as an In Vitro Model to Evaluate Inner Blood-Retinal Barrier Permeability

This protocol illustrates an in vitro endothelial cell transcytosis assay as a model to evaluate inner blood-retinal barrier permeability by measuring the ability of human retinal microvascular endothelial cells to transport horseradish peroxidase across cells in caveolae-mediated transcellular transport processes.

Endotelcelletranscytoseanalyse som en in vitro-modell for å evaluere indre blod-retinal barrierepermeabilitet

Dysfunction of the blood-retinal barrier (BRB) contributes to the pathophysiology of several vascular eye diseases, often resulting in retinal edema and ...

Semi-Automated Planimetric Quantification of Dental Plaque Using an Intraoral Fluorescence Camera

Dental plaque accumulation is quantified using clinical indices or, otherwise, the planimetric plaque index (PPI),&#160;which measures the relative area of a tooth that is covered by plaque deposits. Compared to clinical indices, the PPI has a higher discriminatory power, but traditional planimetry is a time-consuming analysis, as the plaque-covered and clean tooth areas have to be determined manually for each image using image-processing software. Here, we present a method for the semi-automated planimetric quantification of dental plaque, which allows for the rapid processing of up to 1,000 images simultaneously. The method exploits the enhanced contrast between disclosed plaque, sound tooth surfaces, and soft tissues in fluorescence images acquired with an intraoral camera. Careful execution of the clinical procedures and accurate image acquisition are crucial steps for the successful semi-automated identification of the plaque-covered areas. The method is suitable for planimetry on sound facial and oral tooth surfaces, on most composite resin restorations, and on teeth with orthodontic brackets, but not on metallic restorations. Compared to traditional PPI recordings, semi-automated planimetry considerably reduces the amount of time spent on the analysis, as well as the subjective human input, thus increasing the reproducibility of planimetric measurements.

This study presents a semi-automated digital image analysis procedure for the planimetric quantification of disclosed dental plaque based on images acquired with an intraoral fluorescence camera. The method allows for the rapid and reliable quantification of dental plaque in the research environment.

Semi-automatisert planimetrisk kvantifisering av tannplakk ved bruk av et intraoralt fluorescenskamera

Dental plaque accumulation is quantified using clinical indices or, otherwise, the planimetric plaque index (PPI),&#160;which measures the relative area ...

Forebygging av bakre kapselopasifisering gjennom IOL-rotasjon ved kataraktoperasjoner

Rotating the Intraocular Lens to Prevent Posterior Capsular Opacification in Cataract Surgeries

Posterior capsule opacification (PCO) is a common postoperative complication of extracapsular cataract surgery, which is caused by the proliferation and migration of lens epithelial cells and can affect long-term visual outcomes significantly. The most effective treatment for PCO is neodymium-doped yttrium aluminum garnet (Nd:YAG) laser capsulotomy; however, this treatment is associated with posterior segment complication and can break the stability of capsular bag, affecting the position and function of trifocal or toric intraocular lenses (IOLs). Advances in surgical procedures, IOL design, and pharmacy have reduced the rate of PCO in recent years, concentrating on the inhibition of proliferative lens epithelial cells (LECs). This protocol aimed to clear LECs more thoroughly during phacoemulsification and IOL implantation. The first several steps, including clear corneal incision, continuous circular capsulorhexis, hydrodissection, hydrodelineation, and phacoemulsification, were completed as conventional procedures. After placing the IOL into the capsular bag, rotation of the IOL by at least 360&#176; was performed using an irrigation/aspiration tip or a hook, with slight stress on the posterior capsule. Some residuals occurred in the originally transparent capsular bag after rotation of the IOLs. Then, these materials and the viscoelastic were cleared completely using an irrigation/aspiration system. A clear posterior capsule was observed after the surgery in patients undergoing this method. This method of rotating IOLs is a simple, effective, and safe way to prevent PCO by clearing residual LECs and can be carried out without extra tools or skills.

Forfatter Spotlight: Forbedring av visuelle utfall ved kataraktkirurgi: En ny teknikk for å forhindre bakre kapselopasifisering gjennom IOL-rotasjon 

The present protocol describes removing residual epithelial cells by rotating the intraocular lens in extracapsular cataract surgery without extra tools for preventing posterior capsular opacification.

Rotere den intraokulære linsen for å forhindre bakre kapsulær opasifisering ved kataraktoperasjoner

Posterior capsule opacification (PCO) is a common postoperative complication of extracapsular cataract surgery, which is caused by the proliferation and ...

Automatisert prosessor for produksjon av CAR-T-celler i klinisk skala

Chimeric Antigen Receptor T Cell Manufacturing on an Automated Cell Processor

Chimeric antigen receptor (CAR)-T cells represent a promising immunotherapeutic approach for the treatment of various malignant and non-malignant diseases. CAR-T cells are genetically modified T cells that express a chimeric protein that recognizes and binds to a cell surface target, resulting in the killing of the target cell. Traditional CAR-T cell manufacturing methods are labor-intensive, expensive, and may carry the risk of contamination. The CliniMACS Prodigy, an automated cell processor, allows for manufacturing cell therapy products at a clinical scale in a closed system, minimizing the risk of contamination. Processing occurs semi-automatically under the control of a computer and thus&#160;minimizes human involvement in the process, which saves time and reduces variability and errors.
This manuscript and video describes the T cell transduction (TCT) process for manufacturing CAR-T cells using this processor. The TCT process involves CD4+/CD8+ T cell enrichment, activation, transduction with a viral vector, expansion, and harvest. Using the Activity Matrix, a functionality that allows ordering and timing of these steps, the TCT process can be customized extensively. We provide a walk-through of CAR-T cell manufacturing in compliance with current Good Manufacturing Practice&#160;(cGMP) and discuss required release testing and preclinical experiments that will support an Investigational New Drug (IND) application. We demonstrate&#160;the feasibility and discuss the advantages and disadvantages of using a&#160;semi-automatic process for clinical CAR-T cell manufacturing. Finally, we describe an ongoing investigator-initiated clinical trial that targets pediatric B-cell malignancies [NCT05480449] as an example of how this manufacturing process can be applied in a clinical setting.

Forfatter Spotlight: Fremskritt innen CAR-T-celleproduksjon og genterapiproduksjon 

This article details the manufacturing process for chimeric antigen receptor T cells for clinical use, specifically using an automated cell processor capable of performing viral transduction and cultivation of T cells. We provide recommendations and describe pitfalls that should be considered during the process development and implementation of an early-phase clinical trial.