Studies on cationic ocular emulsions containing bipartitioned oil droplets to codeliver cyclosporin A and etodolac.

Background: To prepare ocular emulsions containing bipartitioned oil droplets to entrap cyclosporin A (0.05% w/w) and etodolac (0.2% w/w) by using castor, olive and silicon oils. Methods: The physicochemical characterizations of prepared emulsions were performed. The drug's biodistribution profiles and pharmacokinetic parameters from emulsions were checked using the ultraperformance liquid chromatography-tandem mass spectrometry method in the ocular tissues of the healthy rabbit eye model. Results: The emulsions displayed 365.13 ± 7.21 nm size and 26.45 ± 2.09 mV zeta potential. The ferrying of two drugs after releasing from emulsions occurred across corneal/conjunctival tissues to enter the vitreous and sclera following a single drop administration into the rabbit's eyes. Conclusion: The dual drug-loaded emulsions were more likely to produce synergistic anti-inflammatory activity for managing moderate-to-severe dry eye disease.

Post-chemotherapy changes in retinal pigment epithelium in retinoblastoma eyes.

OBJECTIVE: Histopathological analysis of the retinal pigment epithelial (RPE) changes in retinoblastoma (RB) cases who received pre-surgical chemotherapy. DESIGN: Laboratory-based observational study. METHODS: Five-year analysis was performed to identify Retinoblastoma cases who underwent enucleation after receiving systemic chemotherapy. Grossly, RPE cells were observed in flat preparation in small calottes by staining with fluorescein stain in the raw specimens. They were documented under the objective of compound microscope and compared with hematoxylin and eosin-stained slides in the permanent tissue sections. RESULTS: Out of 51 cases of RB, post-chemotherapy enucleation was performed in 17 cases. Mean age of enucleation was 3.2 years. Endophytic RB (11 cases, 64.71%) was more common than the exophytic variety. Choroidal involvement was noted in 8 cases (47.06%), and optic nerve involvement was seen in 5 cases (29.4%). Focal and diffuse RPE changes were seen in one case each (5.88%). Central RPE cell changes near the cell nucleus were seen in all 17 cases (100%), which were documented by both fluorescein and Hematoxylin and eosin stain (100%). Drusens were observed in 8 cases (47.06%), and RPE proliferations were seen in 3 cases (17.65%). CONCLUSION: The study highlights the characteristic histopathological RPE changes after systemic chemotherapy in RB cases. These changes may be attributable to cell nucleus damage after chemotherapy.

Parasitic Eye Diseases: Nuances in Rapid Identification and Advanced Microscopy.

AIM: To study parasitic eye diseases in a tertiary institute of North-east India by live examination of parasites, rapid staining, and scanning electron microscopy (SEM). METHODS: A 12-year retrospective analysis was performed and all patients diagnosed with ocular parasitic diseases were identified. Examination under a compound microscope, fluorescein staining, and scanning electron microscopy were done. RESULTS: A total of 160 ocular parasitosis cases were identified. The cases for which rapid staining and SEM studies were done included Cysticercosis (n = 18, 11.25%), Hydatidosis (n = 5, 3.13%), Dirofilariasis (n = 5, 3.13%), Thelaziasis (n = 3, 1.87%), and Gnathostomiasis (n = 2, 1.25%). Live examination was performed in 11 cases (6.63%) and 8 cases (4.82%) underwent scanning electron microscopy. . CONCLUSION: Fluorescein staining for identification of parasites and SEM study helped in detailing microscopic and ultrastructural findings.

Histopathology of pigment dispersion syndrome and glaucoma: A Case Report.

Pigment dispersion syndrome (PDS) can have varied manifestations. Anterior and posterior segment involvement by dispersed pigments was documented in gross pathology and by staining under microscopy. Pigmentary changes in the sclera, cornea, anterior chamber, iris, trabecular meshwork, lens, and retinal pigment epithelium and optic nerve findings were consistent with PDS. External scleral and vitreous pigmentation has never been reported before in the scientific literature. Retinal pigment degeneration and granule dispersion were seen throughout the retina which could be a contributing factor in the etiology of PDS.

Intraocular and neuro-cysticercosis with diffuse stromal choroiditis.

Intraocular cysticercosis with central nervous system involvement is not that rare. We report a male child with a right-sided painful blind eye who had intraocular cysticercosis and granuloma in the left frontal lobe of the brain. There was an incidental finding of chronic inflammation in the choroid of that eye supported by histopathology. Immunohistochemistry for T-cells marker and B-cells marker was variable. The patient was treated with antiparasitic, anti-epileptic medications, and oral steroids subsequently.

Studies on spray dried topical ophthalmic emulsions containing cyclosporin A (0.05% w/w): systematic optimization, in vitro preclinical toxicity and in vivo assessments.

Cyclosporin A (CsA, 0.05% w/w)-loaded positively charged emulsions were prepared based on castor oil, chitosan, poloxamer 188, glycerin and double-distilled water. To augment the shelf/storage-stability of original emulsions, the solid-dry powder for reconstitution was made by spray drying technique. The screening (Taguchi OA) and optimization (face-centered central composite) designs produced the optimized conditions for spray drying: 40 Nm3/h aspirator flow rate, 15 ml/min feed rate, 115 °C inlet temperature, 10% mannitol and 1.25% trehalose. The % drug entrapment efficiency values of original and reconstituted emulsions ranged from 73.20 ± 0.13 to 71.55 ± 1.25%. At 20 min post-dissolution, two times higher CsA release was seen from reconstituted emulsions than the original emulsions (85.78 ± 1.14 vs. 42.25 ± 1.84%) in simulated tear fluid. Using MTT assay, the reconstituted emulsions with or without CsA produced 94.512 ± 2.12 to 99.941 ± 1.89% cell viability values in HCE-2 cells. No appreciable change in capillary integrity was visualized in HET CAM following reconstituted emulsions treatment. At equivalent 15 µg drug, the in vitro protein denaturation assay showed augmented inhibition value (~ 85%) for tested CsA emulsions compared to diclofenac reference (68.30 ± 2.05) indicating enhanced anti-inflammatory activity. The CsA concentrations in multiple ocular matrices of rabbit eyes determined by the UPLC-MS/MS method attained the therapeutic drug level of 50-300 ng/ml even at 90 min post-topical instillation of both emulsions. Overall, the CsA emulsion eyedrops can be supplied as a spray dried storable intermediate product for reconstitution.

IgG4-related disease as multiple head and neck swellings: supported by histopathology and immunohistochemistry.

A 66-year-old North-East Indian male presented with bilateral eyelid swelling, ptosis, and bilateral submandibular gland enlargement. Dry skin on both arms was another peculiar complaint. Contrast enhanced CT scans revealed homogenously enhancing, diffusely enlarged lacrimal glands and blood investigations showed raised serum IgG4 levels. Histopathology from lacrimal gland biopsy showed lymphoplasmacytic infiltrates in storiform pattern. Immunohistochemistry showed 35% plasma cells positive for IgG4. A diagnosis of IgG4-related disease was made, due to supportive histopathology, immunohistochemistry, and serum IgG4 levels. The patient showed excellent response to systemic immunomodulators. Abbreviations: IgG4-RD = IgG4-related disease, CECT = Contrast-enhanced computed tomography, ACE = Angiotensin converting enzyme, IHC = Immunohistochemistry, HPF = high power field, IgG4-ROD = IgG4-related ophthalmic disease, ACR/ EULAR = American College of Rheumatology/ European League Against Rheumatism.

Pathologic evidence of retinoblastoma seeds supported by field emission scanning electron microscopy and Raman spectroscopy.

PURPOSE: The aim of this study was to examine the pathology of retinoblastoma (RB) seeds with supportive evidence by field emission scanning electron microscopy and Raman spectroscopy. METHODS: This study was a laboratory-based observational study. Enucleated eyeballs received in the ocular pathology department of a tertiary eye care center in northeast India were included in the cohort after obtaining written informed consent during the surgery. The study was carried out for 6 years (2015-2020). Most of the eyeballs were Group-E RBs. Standard eyeballs sectioning were done by bread loaf techniques. Gross documentations included RB seeds seen in the smallest calotte done with utmost care. Seeds were documented also in permanent sections. Scanning electron microscopy and Raman spectroscopy were carried out in an index case. RESULTS: Out of the total 59 cases, 35 RB cases had different seedings. The mean age at enucleation was 2.9 years. RB seeds were seen in vitreous (n = 19), subretinal plus vitreous (n = 7), anterior chamber (n = 1), over crystalline lens (n = 3), retinal surface (n = 1), retinal pigment epithelium (RPE; n = 2), subretinal (n = 1), calcified seeds (n = 2). Other characteristics were dusts (n = 7), clouds (n = 11), spheres (n = 4), and unspecified type (n = 13). Histopathological high-risk factors showed significant choroidal (n = 22) and optic nerve (n = 15) involvement. Few cases had extraocular spread. Undifferentiated tumor (n = 24) was seen with higher evidence of necrosis (n = 23). Raman spectra differentiated the seeds from the normal tissue on the basis of lipid and protein content. CONCLUSION: This study highlights the different types of RB seeds with high-risk factors. The morphology of those seeds showed the difference between vitreous and subretinal seeds under advanced microscopic observations.

Fluorescein dye as a novel cost-effective approach for staining raw specimens in ophthalmic pathology.

PURPOSE: To study the usefulness of sodium fluorescein dye for staining raw specimens in ophthalmic pathology. METHODS: Laboratory-based observational study. Eye specimens received in the ocular pathology department of a tertiary eye care center in northeast India were included in the study after obtaining the informed consent. The study period was from 2016 to 2019. Specimens received were a corneal button, lid, orbital tissues, enucleated eyeballs, eviscerated eye, explanted intraocular lens (IOLs), optic nerve and ocular parasites. Sections of the gross specimens were stained with sodium fluorescein (C20H12O5NA) dye. The average duration of tissue-stain contact time was 45 s. The sections were analyzed under the compound microscope. The intensity of illumination of the microscope was modulated to obtain high contrast digital photographs. RESULTS: 26 corneal buttons with or without limbal tissue specimens were analyzed with fluorescein staining procedure; limbus with its pigmented cells were seen in the enucleated eyeballs. 33 enucleated eyes (retinoblastoma [RB] (n = 24), phthisical eyes (n = 4), choroidal melanomas (n = 2), and others (n = 3) were included in the study cohort. In these 33 enucleated eyes, vitreous were also examined for the presence of hyalocytes and other pathological cells. Retinal pigment epithelial cells were also seen (n = 11). RB seeds were seen with fluorescein stain and documented in 14 specimens. The RB seeds were mostly in vitreous (n = 9) and subretinal space (n = 5). Fat cells (n = 8) from orbital tissues and sebaceous cells (n = 5) from frozen section specimens were also observed and documented. CONCLUSION: This study highlights a novel method of rapid staining of gross ophthalmic pathology specimens.

Idiopathic inflammatory diseases of orbit and ocular adnexa: Histopathological and immunochemical analysis.

Purpose: : To present histopathological and immunohistochemical analysis of idiopathic inflammatory diseases of orbit and ocular adnexa. Methods: Design- A retrospective laboratory-based study. The study was carried out in an ocular pathology laboratory in a tertiary institute of northeast India where analysis of 93 cases was done in 5 years, during the period from 2011 to 2016. Hematoxylin--eosin and special stains were done for the diagnoses. Immunohistochemistry (IHC) panel was also carried out. For infectious pathology, Grocott's methenamine silver (GMS) stain for fungus, tissue Gram's stain for bacteria's, and acid-fast stains for tubercular bacilli were done. IHC panels were done for CD 20 (B-cells), CD-3 (T-cells), CD-45 (Leukocyte common antigen, LCA), BCL-2, CD-138 (Plasma cells), Kappa, Lambda, IgG-4 in tissue, IgG-4 in serum, etc. IHCs were done using kit methods (standardized) and adequate controls were taken for each sample. Results: 93 cases of nonspecific orbital inflammation were reported out of 1,467 specimens. Orbital pseudotumors (idiopathic orbital inflammatory disease, IOID) were seen in 27 cases (sclerosing variety-6); benign lymphoid hyperplasia in two cases; reactive lymphoid hyperplasia in 10 cases; atypical plasma lymphoproliferative reactive (polyclonal immunophenotypically, IgG4 negative) lesions in four cases; IgG-4 related disease in one case; nonspecific inflammatory reactions (conjunctiva, sclera, and lid) in 49 cases. In all the diagnoses, infections and lymphomas were excluded. Conclusion: Biopsy supported study on nonspecific orbital inflammation was important to know the pattern.

Human ocular thelaziasis caused by gravid Thelazia callipaeda - A unique and rare case report.

An 80-year-old male reported to the clinic with complaints of diminished vision, foreign body sensation, and occasionally some black object moving in front of the right eye. Detailed ocular examination revealed three slender creamy white live worms in the conjunctival sac of the right eye. Total three worms were retrieved and sent to the pathology department for detailed examination. The worm was identified as gravid Thelazia callipaeda. As per the search results in PubMed and Cochrane search engine, this is the first report of human case of gravid Thelazia infestation with simultaneous existence of embryonated or ensheathed eggs and primary-stage larvae in the same worm.

Immunohistochemistry on pattern of ocular & adnexal tumours in a tertiary eye care centre of Northeast India.

BACKGROUND & OBJECTIVES: Ocular and adnexal tumours are important causes of morbidity in India and globally. Immunohistochemistry (IHC) is a vital molecular pathology tool, which helps to diagnose a tumour with more accuracy. The present study was undertaken to document the profile of ocular and adnexal tumour with IHC at a tertiary eye care center in Northeast India. METHODS: This was a prospective and laboratory-based study. Histopathological and IHC study of the ocular and adnexal tumour was carried out from 2012 to 2014. Selection of pathological cases was made on the result of the histological diagnosis. All samples were subjected to IHC using kits for different antibodies as per indications. RESULTS: In total, 645 tumours were included in our study, with 449 benign conditions and 196 were malignant tumours. Total IHCs were done in 87 tumours and 238 of antibodies were used. Non-Hodgkin's lymphomas (B-cell, low-to-intermediate type and mucosal-associated lymphoid tumours) were the most common tumor. INTERPRETATION & CONCLUSIONS: Clinical utility of the IHCs in different ophthalmic tumours can enable pathologists to make an accurate diagnosis and thus help in the overall management of the patient care. IHC may be carried out using various methods and some of the methods practiced are time consuming and tedious. In this study, kit methods were used which were found to be simpler and less time-consuming.

Systemic diffuse large B-cell lymphoma masquerading as neovascular glaucoma.

We describe a case of spontaneous hyphema associated with anterior uveitis presents in a 69-year old female as the prominent sign of the intraocular spread of systemic diffuse large B-cell lymphoma (DLBCL). She had a history of diabetes and initially misdiagnosed as neovascular glaucoma. Clinical history of systemic lymphoma, characteristic findings on B-scan ultrasonography and magnetic resonance imaging scan, and identification of atypical lymphoid cells in aqueous sample established the diagnosis of intraocular metastasis of systemic DLBCL. Therefore, this report highlights that life-threatening malignant systemic lymphoma may masquerade as anterior segment ocular inflammation or neovascular glaucoma.

Molecular pathologic interpretation of new retinoblastoma rosettes.

Newly described retinoblastoma (RB) rosettes have various kinds of pathological significance. Some of their characteristics have been revealed in this immunohistochemistry study. Five paraffin-embedded eyeballs with RB and new rosettes were studied for neuron-specific enolase (NSE), p53, p16, BAX, c-Myc, glial fibrillary acidic protein, synaptophysin, and chromogranin. They were compared and interpreted using control specimens. NSE, P53, and P16 were significantly expressed in the cells of the new rosettes. The presence of new RB rosettes that mostly have histopathological high-risk factors and p53 positivity may be a strong marker of poor prognosis of RB.

Update on pathology of ocular parasitic disease.

Parasites are a group of eukaryotic organisms that may be free-living or form a symbiotic or parasitic relationship with the hosts. Consisting of over 800,000 recognized species, parasites may be unicellular (Protozoa) or multicellular (helminths and arthropods). The association of parasites with human population started long before the emergence of civilization. Parasitic zoonotic diseases are prevalent worldwide including India. Appropriate epidemiological data are lacking on existing zoonotic parasitic diseases, and newer diseases are emerging in our scenario. Systemic diseases such as cysticercosis, paragonimiasis, hydatidosis, and toxoplasmosis are fairly common. Acquired Toxoplasma infections are rising in immune-deficient individuals. Amongst the ocular parasitic diseases, various protozoas such as Cystoidea, trematodes, tissue flagellates, sporozoas etc. affect humans in general and eyes in particular, in different parts of the world. These zoonoses seem to be a real health related problem globally. Recent intensification of research throughout the world has led to specialization in biological fields, creating a conducive situation for researchers interested in this subject. The basics of parasitology lie in morphology, pathology, and with recent updates in molecular parasitology, the scope has extended further. The current review is to address the recent update in ophthalmic parasites with special reference to pathology and give a glimpse of further research in this field.

An interesting case of angiogenesis in cavernous hemangioma.

Cavernous hemangioma is the most common orbital tumor in adult. There is lot of literatures for clinicopathological features of this tumor. These tumors had been studied for the model of angiogenesis in many of the experimental setups. We present a case of 34-year-old male with this tumor in the left eye with computerized tomography evidence. Postsurgical laboratory findings gave interesting evidence of tumor angiogenesis with tumor endothelial cells and sprouting of the small vessels endothelial cells. Podosome rosette could be conceptualized from the characteristic patterns seen in the tumor.

IgG4-related intraocular inflammation masquerading as ciliary body melanoma in a young girl.

Immunoglobulin G4 (IgG4-related diseases) affects various tissues and organs of the human body. Orbital, adnexal, and scleral inflammations were already reported in the medical literature. To the best of our knowledge, we report the first case of intraocular IgG4-associated inflammatory mass in the ciliary body mimicking as a melanoma in a 23-year-old female from Northeast India. Characteristic histopathology, immunohistochemistry in the tissue, protein chemistry, and raised serum IgG4 were supportive for the diagnosis. As this newly diagnosed disease has multi-organ affection and little is known about its pathogenesis particularly in eye and adnexa, the present case will open many challenges in clinico-pathological diagnosis and research in the future.