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1.
J Hum Genet ; 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38956221

RESUMO

Retinoblastoma (RB) is a childhood retinal neoplasm and commonly treated with cytotoxic chemotherapeutic agents. However, these therapeutic approaches often lead to diverse adverse effects. A precise molecular therapy will alleviate these side effects and offer better treatment outcomes. Over the years, kinases have become potential drug targets in cancer therapy. Hence, we aimed to investigate genetic alterations of putative kinase drug targets in RB. Targeted exome sequencing was performed on 35 RB tumors with paired blood samples using a gene panel consisting of 29 FDA-approved kinase genes. Single nucleotide variants were analyzed for pathogenicity using an in-house pipeline and copy number variations (CNVs) were detected by a depth of coverage and CNVPanelizer. The correlation between genetic changes and clinicopathological features was assessed using GraphPad Prism. Three somatic mutations, two in ERBB4 and one in EGFR were identified. Two of these mutations (ERBB4 c.C3836A & EGFR c.A1196T) were not reported earlier. CNV analysis revealed recurrent gains of ALK, MAP2K2, SRC, STK11, and FGFR3 as well as frequent losses of ATM, PI3KCA and ERBB4. Notably, nonresponsive tumors had a higher incidence of amplifications in clinically actionable genes such as ALK. Moreover, ALK gain and ATM loss were strongly correlated with optic nerve head invasion. In conclusion, our study revealed genetic alterations of druggable kinases in RB, providing preliminary insights for the exploration of kinase-targeted therapy in RB.

2.
Med Oncol ; 41(7): 168, 2024 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-38834895

RESUMO

Retinoblastoma (RB) is a pediatric cancer of the eye that occurs in 1/15000 live births worldwide. Albeit RB is initiated by the inactivation of RB1 gene, the disease progression relies largely on transcriptional alterations. Therefore, evaluating gene expression is vital to unveil the therapeutic targets in RB management. In this study, we employed an RT2 Profiler™ PCR array for a focused analysis of 84 cancer-specific genes in RB. An interaction network was built with gene expression data to identify the dysregulated pathways in RB. The key transcript alterations identified in 13 tumors by RT2 Profiler™ PCR array was further validated in 15 tumors by independent RT-qPCR. Out of 84 cancer-specific genes, 68 were dysregulated in RB tumors. Among the 68 genes, 23 were chosen for further analysis based on statistical significance and abundance across multiple tumors. Pathway analysis of altered genes showed the frequent perturbations of cell cycle, angiogenesis and apoptotic pathways in RB. Notably, upregulation of MCM2, MKI67, PGF, WEE1, CDC20 and downregulation of COX5A were found in all the tumors. Western blot confirmed the dysregulation of identified targets at protein levels as well. These alterations were more prominent in invasive RB, correlating with the disease pathogenesis. Our molecular analysis thus identified the potential therapeutic targets for improving retinoblastoma treatment. We also suggest that PCR array can be used as a tool for rapid and cost-effective gene expression analysis.


Assuntos
Neoplasias da Retina , Retinoblastoma , Retinoblastoma/genética , Retinoblastoma/patologia , Retinoblastoma/metabolismo , Humanos , Neoplasias da Retina/genética , Neoplasias da Retina/patologia , Neoplasias da Retina/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica
3.
Cells ; 12(23)2023 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-38067155

RESUMO

The anterior lens epithelium has the ability to differentiate into lens fibres throughout its life. The present study aims to identify and functionally characterize the adult stem cells in the human lens epithelium. Whole mounts of lens epithelium from donor eyes (normal/cataract) were immunostained for SOX2, gap junction protein alpha 1 (GJA1), PAX6, α, ß and γ-crystallins, followed by a confocal analysis. The functional property of adult stem cells was analysed by their sphere forming ability using cultured lens epithelial cells from different zones. Based on marker expression, the lens epithelium was divided into four zones: the central zone, characterized by a small population of PAX6+, GJA1-, ß-crystallin- and γ-crystallin- cells; the germinative zone, characterized by PAX6+, GJA1+, ß-crystallin- and γ-crystallin-; the transitional zone, characterized by PAX6+, GJA1+, ß-crystallin+ and γ-crystallin-; and the equatorial zone, characterized by PAX6+/-, GJA1+, ß-crystallin+, and γ-crystallin+ cells. The putative lens epithelial stem cells identified as SOX2+ and GJA1 membrane expression negative cells were located only in the central zone (1.89 ± 0.84%). Compared to the other zones, a significant percentage of spheres were identified in the central zone (1.68 ± 1.04%), consistent with the location of the putative adult lens epithelial stem cells. In the cataractous lens, an absence of SOX2 expression and a significant reduction in sphere forming ability (0.33 ± 0.11%) were observed in the central zone. The above findings confirmed the presence of putative stem cells in the central zone of the adult human lens epithelium and indicated their probable association with cataract development.


Assuntos
Catarata , gama-Cristalinas , Adulto , Humanos , gama-Cristalinas/metabolismo , Células Epiteliais/metabolismo , Catarata/metabolismo , beta-Cristalinas/metabolismo , Células-Tronco/metabolismo
4.
Genes (Basel) ; 14(11)2023 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-38002955

RESUMO

Glucocorticoids (GCs) are known to regulate several physiological processes and are the mainstay in the management of inflammatory eye diseases. The long-term use of GC causes raised intraocular pressure (IOP) or ocular hypertension (OHT) in about 30-50% of the susceptible individuals depending on the route of administration, and can lead to steroid-induced secondary glaucoma. The present study aims to understand the role of microRNAs (miRNAs) in differential glucocorticoid (GC) responsiveness in human trabecular meshwork (HTM) cells using small RNA sequencing. The human organ-cultured anterior segment (HOCAS) model was used to identify whether donor eyes were from GC-responders (GC-R; n = 4) or GC-non-responders (GC-NR; n = 4) following treatment with either 100 nM dexamethasone (DEX) or ethanol (ETH) for 7 days. The total RNA was extracted from cultured HTM cells with known GC responsiveness, and the differentially expressed miRNAs (DEMIRs) were compared among the following five groups: Group #1: ETH vs. DEX-treated GC-R; #2: ETH vs. DEX-treated GC-NR; #3: overlapping DEGs between Group #1 and #2; #4: Unique DEMIRs of GC-R; #5: Unique DEMIRs of GC-NR; and validated by RT-qPCR. There were 13 and 21 DEMIRs identified in Group #1 and Group #2, respectively. Seven miRNAs were common miRNAs dysregulated in both GC-R and GC-NR (Group #3). This analysis allowed the identification of DEMIRs that were unique to GC-R (6 miRNAs) and GC-NR (14 miRNAs) HTM cells, respectively. Ingenuity Pathway Analysis identified enriched pathways and biological processes associated with differential GC responsiveness in HTM cells. This is the first study to reveal a unique miRNA signature between GC-R and GC-NR HTM cells, which raises the possibility of developing new molecular targets for the management of steroid-OHT/glaucoma.


Assuntos
Glaucoma , MicroRNAs , Hipertensão Ocular , Humanos , Glucocorticoides/farmacologia , Malha Trabecular/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Hipertensão Ocular/induzido quimicamente , Hipertensão Ocular/metabolismo , Glaucoma/genética , Dexametasona/farmacologia , Análise de Sequência de RNA , Esteroides/metabolismo
7.
Genes (Basel) ; 13(5)2022 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-35627267

RESUMO

Aim: To investigate genes and pathways involved in differential glucocorticoid (GC) responsiveness in human trabecular meshwork (HTM) cells using RNA sequencing. Methods: Using paired human donor eyes, human organ-cultured anterior segment (HOCAS) was established in one eye to characterize GC responsiveness based on intra ocular pressure (IOP) change and, in the other eye, primary HTM cell culture was established. For RNA sequencing, total RNA extracted from GC-responder (GC-R) and non-responder (GC-NR) cells after dexamethasone (DEX) or ethanol (ETH) treatment for 7d was used. Differentially expressed genes (DEGs) were compared among five groups and validated. Results: In total, 616 and 216 genes were identified as significantly dysregulated in Group #1 and #2 (#1: ETH vs. DEX-treated GC-R; #2: ETH vs. DEX-treated GC-NR), respectively. Around 80 genes were commonly dysregulated in Group #3 (overlapping DEGs between #1 and #2), whereas 536 and 136 genes were uniquely expressed in GC-R (#4) and GC-NR HTM (#5) cells, respectively. Pathway analysis revealed that WNT signaling, drug metabolism cytochrome p450, cell adhesion, TGF-ß signaling, and MAPK signaling were associated with GC responsiveness. Conclusion: This is the first study reporting distinct gene signatures and their associated pathways for GC-R and GC-NR HTM cells. WNT and MAPK signaling are potential therapeutic targets for the management of GC-induced glaucoma.


Assuntos
Glucocorticoides , Malha Trabecular , Perfilação da Expressão Gênica , Glucocorticoides/metabolismo , Glucocorticoides/farmacologia , Humanos , Pressão Intraocular , Malha Trabecular/metabolismo , Transcriptoma/genética
8.
Sci Rep ; 12(1): 8299, 2022 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-35585182

RESUMO

In the quest of identifying newer molecular targets for the management of glucocorticoid-induced ocular hypertension (GC-OHT) and glaucoma (GCG), several microarray studies have attempted to investigate the genome-wide transcriptome profiling of primary human trabecular meshwork (TM) cells in response to dexamethasone (DEX). However, no studies are reported so far to demonstrate the temporal changes in the expression of genes in the cultured human TM cells in response to DEX treatment. Therefore, in the present study, the time-dependent changes in the genome-wide expression of genes in primary human TM cells after short (16 hours: 16 h) and long exposure (7 days: 7 d) of DEX was investigated using RNA sequencing. There were 199 (118 up-regulated; 81 down-regulated) and 525 (119 up-regulated; 406 down-regulated) DEGs in 16 h and 7 d treatment groups respectively. The unique genes identified in 16 h and 7 d treatment groups were 152 and 478 respectively. This study found a distinct gene signature and pathways between two treatment regimes. Longer exposure of DEX treatment showed a dys-regulation of Wnt and Rap1 signaling and so highlighted potential therapeutic targets for pharmacological management of GC-OHT/glaucoma.


Assuntos
Glaucoma , Malha Trabecular , Células Cultivadas , Dexametasona/efeitos adversos , Glaucoma/induzido quimicamente , Glaucoma/tratamento farmacológico , Glaucoma/genética , Glucocorticoides/metabolismo , Humanos , Malha Trabecular/metabolismo , Transcriptoma
9.
Indian J Ophthalmol ; 69(8): 2111-2115, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34304188

RESUMO

Purpose: This study is aimed to investigate the presence of Human papillomavirus (HPV) DNA in tumors obtained from sporadic retinoblastoma patients. Methods: One hundred six tumor tissues obtained from sporadic RB patients were analyzed for HPV infection by use of both seminested PCR and real-time quantitative PCR. Results: Of 106 RB patients, 55 were male and 51 were female. The mean age at diagnosis was 26.77 ± 15.36 (mean ± Std. dev) months. Almost all patients presented with leukocoria. Molecular investigation by different methods revealed no HPV positivity in any tumor genome. Conclusion: Our study demonstrates no association between HPV and RB, postulating HPV may not be a major risk factor in the etiology of RB.


Assuntos
Alphapapillomavirus , Infecções por Papillomavirus , Neoplasias da Retina , Retinoblastoma , Feminino , Humanos , Índia/epidemiologia , Masculino , Papillomaviridae/genética , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/epidemiologia , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias da Retina/diagnóstico , Neoplasias da Retina/epidemiologia , Retinoblastoma/diagnóstico , Retinoblastoma/epidemiologia
10.
Life Sci ; 280: 119750, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34171378

RESUMO

The tumor microenvironment (TME) constitutes multiple cell types including cancerous and non-cancerous cells. The intercellular communication between these cells through TME derived exosomes may either enhance or suppress the tumorigenic processes. The tumor-derived exosomes could convert an anti-tumor environment into a pro-tumor environment by inducing the differentiation of stromal cells into tumor-associated cells. The exosomes from tumor-associated stromal cells reciprocally trigger epithelial-to-mesenchymal transition (EMT) in tumor cells, which impose therapeutic resistance and metastasis. It is well known that these exosomes contain the signals of EMT, but how these signals execute chemoresistance and metastasis in tumors remains elusive. Understanding the significance and molecular signatures of exosomes transmitting EMT signals would aid in developing appropriate methods of inhibiting them. In this review, we focus on molecular signatures of exosomes that shuttle between cancer cells and their stromal populations in TME to explicate their impact on therapeutic resistance and metastasis through EMT. Especially Wnt signaling is found to be involved in multiple ways of exosomal transport and hence we decipher the biomolecules of Wnt signaling trafficked through exosomes and their potential in serving as therapeutic targets.


Assuntos
Transição Epitelial-Mesenquimal , Exossomos/patologia , Neoplasias/patologia , Microambiente Tumoral , Animais , Antineoplásicos/farmacologia , Carcinogênese/efeitos dos fármacos , Carcinogênese/metabolismo , Carcinogênese/patologia , Comunicação Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Exossomos/efeitos dos fármacos , Exossomos/metabolismo , Humanos , Metástase Neoplásica/tratamento farmacológico , Metástase Neoplásica/patologia , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Microambiente Tumoral/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos
11.
Sci Rep ; 11(1): 605, 2021 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-33436790

RESUMO

The purpose of the present study was to assess the differential intraocular pressure response (IOP) to dexamethasone (DEX) treatment at two dose levels (100 or 500 nM) in perfusion cultured Indian cadaveric eyes to investigate glucocorticoid (GC) responsiveness. In a human organ-cultured anterior segment (HOCAS) set-up, the eye pressure was monitored for every 24 h post DEX infusion (100 or 500 nM) or 0.1% ethanol treatment for 7 days after baseline stabilization. The expression of DEX-inducible proteins such as myocilin and fibronectin in HOCAS-TM tissues was assessed by immunostaining. Elevated IOP was observed in 6/16 eyes [Mean ± SEM (mΔIOP): 15.50 ± 1.96 mmHg; 37.5% responders] and 3/15 eyes (Mean ± SEM mΔIOP: 10 ± 0.84 mmHg; 20% responders) in 100 nM and 500 nM dose groups respectively. Elevated IOP in GC responder eyes was substantiated with a significant increase in myocilin (11.8-fold; p = 0.0002) and fibronectin (eightfold; p = 0.04) expression as compared to vehicle-treated eyes by immunofluorescence analysis. This is the first study reporting the GC responsiveness in Indian cadaveric eyes. The observed GC response rate was comparable with the previous studies and hence, this model will enable us to investigate the relationship between differential gene expression and individual GC responsiveness in our population.


Assuntos
Dexametasona/farmacologia , Olho/fisiopatologia , Glaucoma/fisiopatologia , Glucocorticoides/farmacologia , Malha Trabecular/efeitos dos fármacos , Idoso , Cadáver , Células Cultivadas , Olho/efeitos dos fármacos , Glaucoma/tratamento farmacológico , Humanos , Pressão Intraocular , Perfusão
13.
Sci Rep ; 10(1): 10202, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32576873

RESUMO

The intraocular pressure lowering property of a new rho kinase inhibitor, SB772077B (SB77) has been previously demonstrated in perfused human cadaveric eyes. In this study, the efficacy of SB77 in alleviating the aqueous outflow resistance mediated by cyclic mechanical stress in perfused human cadaveric eyes was investigated. A human anterior segment perfusion culture model was used to investigate the effect of cyclic intraocular pressure (IOP) on aqueous outflow facility in presence or absence of SB77. The status of RhoA activation and the downstream effector molecule myosin-light chain phosphorylation (p-MLC) was investigated by Western blot. Cyclic mechanical stress resulted in decrease in aqueous outflow facility (-19.79 ± 4.93%; p = 0.019) in perfused human eyes and treatment with SB77 (50 µM) significantly enhanced outflow facility by 15% (p = 0.05). The increase in outflow facility by SB77 was confirmed with the inactivation of RhoA/ROCK signaling and decreased expression of extracellular matrix markers. SB77 effectively reduced the outflow resistance mediated by cyclic IOP and thus may be a potential clinical candidate for the management of glaucoma.


Assuntos
Humor Aquoso/efeitos dos fármacos , Olho/efeitos dos fármacos , Pressão Intraocular/efeitos dos fármacos , Inibidores de Proteínas Quinases/uso terapêutico , Actinas/metabolismo , Idoso , Animais , Cadáver , Matriz Extracelular/metabolismo , Olho/metabolismo , Humanos , Cadeias Leves de Miosina/metabolismo , Técnicas de Cultura de Órgãos/métodos , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Estresse Mecânico , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo
14.
Curr Cancer Drug Targets ; 20(9): 710-719, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32364077

RESUMO

BACKGROUND: Retinoblastoma is a sight and life-threatening embryonal tumor in children. Though chemotherapy is the main mode of therapy, evolving resistance remains a major obstacle in treatment success. The presence of cancer stem cells (CSC) is frequently reported to be responsible for chemoresistance in multiple tumors. OBJECTIVE: Our study aims to identify the molecular factors that facilitate the chemoresistance through cancer stem cells in retinoblastoma. METHODS: We developed etoposide and carboplatin resistant retinoblastoma (Y79) cell lines by stepwise drug increment treatment, validated with MTT and TUNEL assays. Colony forming and invasive ability were studied by soft-agar colony forming and transwell assays, respectively. Similar analysis in non-responsive retinoblastoma tumors were carried out by histopathology. Finally, expression of CSC/neuronal markers and ABC transporters were examined by quantitative PCR and protein expression of neuronal stem cell markers was confirmed by Western blot. RESULTS: Larger colony size of resistant cells in soft-agar assay provided evidence for increased selfrenewability. Histopathology in non-responsive tumors showed poorly differentiated cells predominantly. Besides, both resistant cell lines and non-responsive tumors showed increased invasion with higher expression of neuronal stem cell markers - SOX2, NANOG, OCT4 and ABC transporters - ABCB1 and ABCC3. Increased self-renewal ability and invasion along with overexpression of stemness markers in resistant cells and tumors provide evidence for stemness driving chemoresistance and invasion in retinoblastoma. CONCLUSION: We have demonstrated Neuronal stem cell/CSC markers that facilitate the maintenance of cancer stem cells. Developing therapies targeting these factors will help in overcoming resistance and improving retinoblastoma treatment.


Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Carboplatina/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Etoposídeo/uso terapêutico , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neurais/metabolismo , Neoplasias da Retina/tratamento farmacológico , Neoplasias da Retina/metabolismo , Retinoblastoma/tratamento farmacológico , Retinoblastoma/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Carboplatina/farmacologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Etoposídeo/farmacologia , Humanos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Proteína Homeobox Nanog/metabolismo , Invasividade Neoplásica , Fator 3 de Transcrição de Octâmero , Neoplasias da Retina/patologia , Retinoblastoma/patologia , Fatores de Transcrição SOXB1/metabolismo , Transdução de Sinais
15.
Indian J Ophthalmol ; 67(3): 436-439, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30777982

RESUMO

Retinoblastoma has an increased inheritance risk of germline RB1 mutations in offspring and siblings, especially twins. Three families, each having one retinoblastoma-affected twin, were selected for genetic analysis and DNA profiling. Germline RB1 mutations were found in all probands. DNA profiling carried on similar-looking twins of families I and II, proved them to be fraternal. This study demonstrates the importance of genetic analysis of RB1 gene for risk prediction in retinoblastoma families. It also emphasizes that DNA profiling is a mandate for genetic screening of families with twins, thus adding a new dimension in counseling of retinoblastoma.


Assuntos
Doenças em Gêmeos , Testes Genéticos/métodos , Neoplasias da Retina/diagnóstico , Retinoblastoma/diagnóstico , DNA/genética , Feminino , Genes do Retinoblastoma/genética , Mutação em Linhagem Germinativa , Humanos , Lactente , Microscopia Acústica , Linhagem , Neoplasias da Retina/genética , Retinoblastoma/genética
16.
Eye (Lond) ; 33(6): 896-901, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30635644

RESUMO

PURPOSE: To improve and validate the smartphone-based leukocoria detection application so that non-ophthalmologists could make use of the smartphone for early detection of Retinoblastoma (RB) in young children without anesthesia and pharmacological dilatation of the pupil. METHODS: Two apps, MDEyeCare and CRADLE, developed for red reflex based leukocoria detection were used in iPhone 6s. MDEyeCare methodology was modified with respect to ambient lighting, the distance between camera and eye and different gazes for better performance. We analyzed totally 34 eyes of 23 RB patients and four normal children. Each of the RB patients was confirmed with clinical examination and radiological investigations. RESULTS: Modification in the methodology of MDEyeCare app could detect the leukocoria in early stages of RB (50% of Group B, 83% of Group C). In late stages (Group D and E), 100% of tumors were detected. The CRADLE app failed to provide adequate leukocoria detection except four late stage RB eyes. Among the 14 normal eyes (6 from unilateral RB and eight from normal children), pseudo-leukocoria was observed in three eyes only at lateral gaze even with MDEyeCare app. CONCLUSION: Improved methodology in smartphone-based app enhanced the detection of RB and this may translate into better outcome after treatment with respect to vision salvage.


Assuntos
Técnicas de Diagnóstico Oftalmológico/instrumentação , Neoplasias da Retina/diagnóstico , Retinoblastoma/diagnóstico , Smartphone , Software , Pré-Escolar , Feminino , Seguimentos , Humanos , Imageamento por Ressonância Magnética , Masculino , Reprodutibilidade dos Testes , Estudos Retrospectivos
17.
Sci Rep ; 8(1): 15472, 2018 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-30341380

RESUMO

We investigated the effect of a new Rho kinase inhibitor, SB772077B (SB77) on aqueous outflow facility (OF) in human eyes using human organ-cultured anterior segment (HOCAS). IOP was monitored for 24 h post-treatment with either SB77 (0.1/10/50 µM) or vehicle after a stable baseline pressure. The hydrodynamic pattern of aqueous outflow was analysed by labelling outflow pathway with red fluorescent microspheres. The effect of SB77 on cell morphology, actin stress fibers, focal adhesions, ECM, status of RhoA activation and myosin light chain phosphorylation (p-MLC) were evaluated and compared with Y27632, by immunostaining using primary human trabecular meshwork (HTM) cells. Following 24 h treatment, SB77 increased OF by 16% at 0.1 µM (N = 6), 29% at 10 µM (N = 8; p = 0.018) and 39% at 50 µM (N = 8; p = 0.004) in human eyes. There was an overall increase in tracer quantity and in area along inner wall of Schlemm's canal. Treatment with SB77 showed no evidence of cytotoxicity and caused a significant reduction in the expression of fibrotic markers compared to Y27632. The present findings indicate that SB77 treatment was effective in enhancing OF and reducing fibrotic markers in an ex vivo model. Thus SB77 may be a potential clinical candidate for the management of glaucoma.


Assuntos
Humor Aquoso/metabolismo , Inibidores Enzimáticos/metabolismo , Olho/efeitos dos fármacos , Quinases Associadas a rho/antagonistas & inibidores , Humanos , Hidrodinâmica , Modelos Biológicos , Técnicas de Cultura de Órgãos
18.
Indian J Ophthalmol ; 63(7): 600-5, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26458478

RESUMO

PURPOSE: The purpose of this study is to develop methods to identify glaucoma by examining the optic nerve head (ONH) of donor's eyes when information on the preexisting ocular disease is unavailable. MATERIALS AND METHODS: The ONH of the donor's eyes was evaluated under a stereomicroscope for the cup-disc ratio (CDR) and focal retinal rim thinning. The vertical diameter of the cup and disc was also measured using a precalibrated eyepiece micrometer. The suspect eyes were subjected to histological analysis to confirm the presence of specific glaucomatous changes. RESULTS: A total of 202 eyes from 119 donors (68 males and 51 females, aged 42-96) were evaluated for glaucoma. Among them, 190 (94%) eyes showing vertical CDR in the of 0.0-0.6 range were considered nonglaucomatous and the remaining eyes with >0.6 as glaucoma suspect. The calculated mean CDR of the two groups (0.3 ± 0.16, 0.62 ± 0.27) was highly significant (P = 0.0003). Of 12 eyes suspected of glaucoma, 7 eyes from 5 donors showed specific glaucomatous changes by histology. The prevalence of glaucoma was 4.2% among the donors studied. CONCLUSIONS: A simple method of screening fresh donor eyes for selecting those with glaucoma features using CDR and histological analysis was reported. This method helps to obtain biologically active human ocular tissue for glaucoma research on gene expression, ultrastructural/proteome changes, and outflow mechanism.


Assuntos
Bancos de Olhos , Glaucoma de Ângulo Aberto/complicações , Pressão Intraocular , Disco Óptico/patologia , Doenças do Nervo Óptico/diagnóstico , Doadores de Tecidos , Campos Visuais/fisiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Glaucoma de Ângulo Aberto/diagnóstico , Glaucoma de Ângulo Aberto/epidemiologia , Humanos , Índia/epidemiologia , Masculino , Pessoa de Meia-Idade , Doenças do Nervo Óptico/etiologia , Prevalência , Tomografia de Coerência Óptica , Testes de Campo Visual
19.
Asia Pac J Clin Oncol ; 11(4): 299-307, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26264229

RESUMO

AIM: Retinoblastoma (RB) is the most common primary intraocular malignancy affecting children under 5 years of age. This study aims to correlate the clinical parameters with RB1 mutation in the light of Knudson's two-hit hypothesis in Indian RB patients. METHODS: We analyzed the clinical details of 73 RB patients visiting Aravind Eye Hospital, Madurai, India, between January and October 2012. Data on gender, presenting age and sign, laterality, number of tumors in each eye and family history were collected. A semi log plot was derived based on Knudson's two-hit hypothesis. Genetic analysis of RB1 was carried out to identify the two hits. RESULTS: The mean age at diagnosis for unilateral and bilateral cases was 24.0 ± 15.1 and 9.8 ± 11.5 months, respectively. Familial RB was seen in 13 (17.8%) patients of whom 11 were bilateral. Multiple tumors were observed more frequently in bilateral than in unilateral cases. All unilateral and bilateral patients followed the two-hit and one-hit curves, respectively, confirming Knudson's hypothesis in Indian patients. Genetic analysis identified two somatic mutations in tumor samples of sporadic unilateral cases. Among the two bilateral patients, one received the first hit from her father and the other patient developed a de novo germline mutation during early development. CONCLUSION: The two-hit hypothesis has been reestablished in Indian patients. Genetic analysis of tumor samples has also complemented the statistical analysis to reaffirm the two hits in tumor development.


Assuntos
Predisposição Genética para Doença , Testes Genéticos , Mutação/genética , Proteína do Retinoblastoma/genética , Retinoblastoma/genética , Criança , Pré-Escolar , Análise Mutacional de DNA , Progressão da Doença , Feminino , Humanos , Índia/epidemiologia , Lactente , Recém-Nascido , Masculino , Retinoblastoma/epidemiologia
20.
Cornea ; 34(11): 1478-86, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26312622

RESUMO

PURPOSE: Using in vivo confocal microscopy, we established that unique hyperreflective structures in the anterior limbal stroma of healthy individuals represent the limbal stromal niche. The aim of this study was to characterize the limbal stromal microarchitecture in patients with limbal stem cell deficiency (LSCD). METHODS: After obtaining informed consent, 10 patients with LSCD and 3 with macular corneal dystrophy were recruited. In vivo confocal imaging of the limbus and cornea of the affected and normal eyes was performed using an HRT III laser scanning microscope, beyond the epithelium deep into the stroma. RESULTS: In the case of LSCD, the limbal epithelium was replaced by conjunctival epithelium. A large number of inflammatory and dendritic cells were identified along with blood vessels from the epithelium to deep stromal layers. The unique hyperreflective niche structures were replaced by homogenously bright fibrous structures in all eyes with total LSCD. In patients with partial LSCD, even the clinically defined normal limbus had fibrotic stroma. In a patient with focal LSCD, only the affected limbal stroma remained fibrotic, whereas the adjacent clinically normal limbus had the unique hyperreflective structures. Although the opaque corneal stroma appeared bright because of proteoglycan deposition, it was possible to identify the normal limbal epithelial and stromal architecture in macular corneal dystrophy. CONCLUSIONS: In the case of LSCD, the limbal stromal niche was replaced by bright fibrotic structures indicating persistence of damage several months after injury. Further studies are required to characterize the sequential events occurring in the anterior limbal stroma after injury using this noninvasive method.


Assuntos
Distrofias Hereditárias da Córnea/patologia , Opacidade da Córnea/patologia , Substância Própria/patologia , Epitélio Corneano/patologia , Limbo da Córnea/patologia , Células-Tronco/patologia , Adulto , Contagem de Células , Substância Própria/citologia , Feminino , Humanos , Limbo da Córnea/citologia , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Transplante de Células-Tronco
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