Analysis of sociodemographic profile of glaucoma patients with risk factors, subtypes, and disease severity in a tertiary eye care facility in Northern India.

PURPOSE: To study the different subtypes of glaucoma and to analyze the demographic and clinical profile of glaucoma patients along with associated risk factors in a multi-locational North Indian study. METHODS: Newly diagnosed and known glaucoma patients on treatment were enrolled in the study. Complete history was taken, and clinical evaluation was done by senior glaucoma experts. RESULTS: Five hundred and eighty-six patients (1121 eyes) were included with 383 males (65.4%) and 203 females (34.6%). Of these, 508 (86.7%) had primary glaucoma and 78 (13.3%) had secondary glaucoma. Also, 263 (44.9%) patients had primary open angle glaucoma (POAG) and 172 (29.4%) had primary angle closure glaucoma. Four hundred and fifty-one (77%) were graduates or postgraduates and 293 (50%) were dependents on family income. One hundred and forty-one (24%) had a positive family history. Secondary glaucoma patients had a lower average age (52.6 ± 16.8 years) and higher average intraocular pressure (IOP; 34.23 ± 13 mm) compared to primary glaucoma patients (26.6 ± 9.7 mm) (P < 0.0001). Thirty-three (47.8%) out of 78 secondary glaucoma patients had advanced disease. Seventy-four (12.6%) were first-time diagnosed and 18 had advanced disease. In addition, 161 eyes had one or more glaucoma surgeries done (128 trabeculectomy and 28 phacotrabeculectomy). Ninety-five (16.2%) patients were advised surgery at our centers (46 trab, 26 phacotrab, 15 phaco, five Ahmed glaucoma valve implantation, and three pars plana vitrectomy). Disease progression was noted in 62 patients reporting good compliance to medicines. CONCLUSION: POAG was the most common variant in this study. Secondary glaucoma patients were youngest in age and had the highest IOP. Trabeculectomy was the most performed surgery. A good literacy rate did not prevent late presentations.

Complex interplay between RAS GTPases and RASSF effectors regulates subcellular localization of YAP.

RAS GTPases bind effectors to convert upstream cues to changes in cellular function. Effectors of classical H/K/NRAS are defined by RBD/RA domains which recognize the GTP-bound conformation of these GTPases, yet the specificity of RBD/RAs for over 160 RAS superfamily proteins remains poorly explored. We have systematically mapped interactions between BRAF and four RASSF effectors, the largest family of RA-containing proteins, with all RAS, RHO and ARF small GTPases. 39 validated complexes reveal plasticity in RASSF binding, while BRAF demonstrates tight specificity for classical H/K/NRAS. Complex between RASSF5 and diverse RAS GTPases at the plasma membrane can activate Hippo signalling and sequester YAP in the cytosol. RASSF8 undergoes liquid-liquid phase separation and resides in YAP-associated membraneless condensates, which also engage several RAS and RHO GTPases. The poorly studied RASSF3 has been identified as a first potential effector of mitochondrial MIRO proteins, and its co-expression with these GTPases impacts mitochondria and peroxisome distribution. These data reveal the complex nature of GTPase-effector interactions and show their systematic elucidation can reveal completely novel and biologically relevant cellular processes.

Sustainable Synthesis of Novel Green-Based Nanoparticles for Therapeutic Interventions and Environmental Remediation.

The advancement in nanotechnology has completely revolutionized various fields, including pharmaceutical sciences, and streamlined the potential therapeutic of many diseases that endanger human life. The synthesis of green nanoparticles by biological processes is an aspect of the newly emerging scientific field known as "green nanotechnology". Due to their safe, eco-friendly, nontoxic nature, green synthesis tools are better suited to produce nanoparticles between 1 and 100 nm. Nanoformulation of different types of nanoparticles has been made possible by using green production techniques and commercially feasible novel precursors, such as seed extracts, algae, and fungi, that act as potent reducing, capping, and stabilizing agents. In addition to this, the biofunctionalization of nanoparticles has also broadened its horizon in the field of environmental remediation and various novel therapeutic innovations including wound healing, antimicrobial, anticancer, and nano biosensing. However, the major challenge pertaining to green nanotechnology is the agglomeration of nanoparticles that may alter the surface topology, which can affect biological physiology, thereby contributing to system toxicity. Therefore, a thorough grasp of nanoparticle toxicity and biocompatibility is required to harness the applications of nanotechnology in therapeutics.

Novel EGFR mutations in diffuse midline gliomas using cost-effective strategies: A report of 2 cases.

Background: Diffuse midline gliomas (DMGs) are malignant tumors predominantly affecting children, often leading to poor outcomes. The 2021 World Health Organization classification identifies 3 subtypes of DMGs, all characterized by the loss of H3K27 trimethylation. Here, we report 2 cases of DMG with Epidermal Growth Factor Receptor (EGFR) mutations within exon 20, contributing to the understanding of the molecular complexity of these pediatric brain tumors. Methods: An economical immunohistochemical panel was designed to aid in the diagnosis of most DMGs in resource-constrained regions. Sanger sequencing was employed to identify rare EGFR mutations in exon 20 of 2 cases. Results: Molecular analyses of 2 cases of DMG revealed novel EGFR mutations within exon 20. These mutations were identified using cost-effective diagnostic approaches. The presence of EGFR mutations expands the molecular landscape of DMGs and highlights the genetic heterogeneity within this tumor entity. Conclusions: These findings underscore the molecular heterogeneity of DMGs and the significance of identifying novel mutations, such as EGFR mutations in exon 20. Further research into the molecular mechanisms underlying DMGs is warranted to advance therapeutic strategies and improve outcomes for pediatric patients.

Developing a model for aqueous deficient dry eye secondary to periglandular cicatrizing conjunctivitis.

PURPOSE: The current study used various techniques to develop a rabbit animal model of lacrimal gland damage caused by scarring conjunctivitis in the periglandular area. METHODS: Left eyes of New Zealand white rabbits were injected with 0.1 ml of 1M NaOH subconjunctivally around superior and inferior lacrimal gland orifices (Group 1, n = 4), touched with 1M NaOH for 100 s to the superior and inferior fornices with conjunctival denuding (Group 2; n = 4), and electrocauterization to the ductal opening area (Group 3; n = 4). The ocular surface staining, Schirmer I, lacrimal gland, and conjunctival changes were observed at baseline,1, 4, 8, and 12 weeks. The degree of glandular inflammation, conjunctival fibrosis (Masson Trichrome), and goblet cell density (PAS) were also assessed. RESULTS: At 12 weeks, the lacrimal glands of group 1 rabbits with periglandular injection showed severe inflammation with mean four foci/10HPF and a significant mean reduction in the Schirmer values by 7.6 mm (P = 0.007). Lacrimal glands had diffuse acinar atrophy, loss of myoepithelial cells, and ductular dilatation. The overlying conjunctiva showed fibrosis, goblet cell loss, and corneal vascularization in the inferotemporal quadrant. No lacrimal gland or ocular surface changes were observed in groups 2 and 3 at 12 weeks, except for localized subconjunctival fibrosis. CONCLUSION: Periglandular injection of 0.1 ml of 1M NaOH induced extensive lacrimal gland damage with reduced secretion and scarring in the subconjunctival plane compared to direct cauterization or direct NaOH contact to the ductal orifices of the rabbit lacrimal gland.

Meibography-Guided Electroepilation for Congenital Distichiasis: A Novel Surgical Technique.

PURPOSE: Congenital distichiasis is managed either by ablation, using laser, cryotherapy, or electroepilation, or by surgical excision with mucous membrane grafting. Ablative procedures are usually blind as the exact depth of distichiatic eyelashes is unknown. The described surgical technique utilizes meibography for imaging the root and depth of distichiatic eyelashes that aided in performing electroepilation. METHODS: Six patients (n = 24 eyelids; mean age 15.5 ± 12.2 years) underwent infrared meibography (Oculus Keratograph 5 M) and noninvasive tear breakup time prospectively. Eyelashes were electroepilated using a premarked needle inserted at a depth based on meibography findings in 4 patients. Surgical success was defined as no distichiatic eyelash regrowth and functional success was defined as the resolution of symptoms at a minimum of 3 months of follow-up. RESULTS: All 6 patients had all 4 eyelids involved to varying degrees, with a total of 230 distichiatic eyelashes. The median number of distichiatic eyelashes was 9 in the upper eyelids and 4.5 in the lower eyelids. Meibography revealed visible distichiatic eyelash roots in 70% of eyelashes in the upper eyelid and 87.8% in the lower eyelid, respectively. The median eyelash root depth was 2.7 mm (mean 2.9 mm, range 1.8-5.4 mm). The mean noninvasive tear breakup time was 12.2 seconds despite absent or rudimentary meibomian gland segments seen on meibography. The anatomical success was 75% (12/16 eyelids), and functional success was 87.5% (7/8 eyes) at a median follow-up of 5.5 months. CONCLUSION: Preoperative infrared meibography in eyelids with congenital distichiasis helps estimate the eyelash depth and can be used to guide eyelash ablation procedures.

Biallelic loss of function variants in FUZ result in an orofaciodigital syndrome.

Orofaciodigital syndrome is a distinctive subtype of skeletal ciliopathies. Disease-causing variants in the genes encoding the CPLANE complex result in a wide variety of skeletal dysplasia with disturbed ciliary functions. The phenotypic spectrum includes orofaciodigital syndrome and short rib polydactyly syndrome. FUZ, as a part of the CPLANE complex, is involved in intraflagellar vesicular trafficking within primary cilia. Previously, the variants, c.98_111+9del and c.851G>T in FUZ were identified in two individuals with a skeletal ciliopathy, manifesting digital anomalies (polydactyly, syndactyly), orofacial cleft, short ribs and cardiac defects. Here, we present two novel variants, c.601G>A and c.625_636del in biallelic state, in two additional subjects exhibiting phenotypic overlap with the previously reported cases. Our findings underscore the association between biallelic loss of function variants in FUZ and skeletal ciliopathy akin to orofaciodigital syndrome.

Two sisters with RSPRY1-related spondyloepimetaphyseal dysplasia.

Biallelic variants in RSPRY1 have been found to result in spondyloepimetaphyseal dysplasia. Two siblings presenting with short stature, facial dysmorphism, progressive vertebral defects, small epiphysis, cupping and fraying of metaphyses, brachydactyly, and short metatarsals harbored a homozygous missense variant c.1652G>A;p.(Cys551Tyr) in the RSPRY1 gene. The phenotype in our patients resembles spondyloepimetaphyseal dysplasia, Faden-Alkuraya type. Thus, our study provides further evidence to support the association of RSPRY1 variants with spondyloepimetaphyseal dysplasia. We observed joint dislocation as a novel clinical feature of this condition.

A gain of function mutation in AKT1 increases hexokinase 2 and diminishes oxidative stress in meningioma.

Increasing evidence suggests the oncogenic role of missense mutation (AKT1-E17K) of AKT1 gene in meningiomas. Upon investigating the connection between the pro-tumorigenic role of AKT1-E17K and cellular metabolic adaptations, elevated levels of glycolytic enzyme hexokinase 2 (HK2) was observed in meningioma patients with AKT1-E17K compared to patients harboring wild-type AKT1. In vitro experiments also suggested higher HK2 levels and its activity in AKT1-E17K cells. Treatment with the conventional drug of choice AZD5363 (a pan AKT inhibitor) enhanced cell death and diminished HK2 levels in AKT1 mutants. Given the role of AKT phosphorylation in eliciting inflammatory responses, we observed increased levels of inflammatory mediators (IL-1ß, IL6, IL8, and TLR4) in AKT1-E17K cells compared to AKT1-WT cells. Treatment with AKT or HK2 inhibitors dampened the heightened levels of inflammatory markers in AKT1-E17K cells. As AKT and HK2 regulates redox homeostasis, diminished ROS generation concomitant with increased levels of NF-E2- related factor 2 (Nrf2) and superoxide dismutase 1 (SOD1) were observed in AKT1-E17K cells. Increased sensitivity of AKT1-E17K cells to AZD5363 in the presence of HK2 inhibitor Lonidamine was reversed upon treatment with ROS inhibitor NAC. By affecting metabolism, inflammation, and redox homeostasis AKT1-E17K confers a survival advantage in meningioma cells. Our findings suggest that targeting AKT-HK2 cross-talk to induce ROS-dependent cell death could be exploited as novel therapeutic approach in meningiomas.

Neuroinflammation and Acetylcholinesterase Inhibitory Potentials of a Spiroketal-Enol Ether Polyyne Isolated from Artemisia pallens Wall. ex DC.

Artemisia pallens Wall. ex DC (Asteraceae) is cultivated for the production of high-value essential oil from its aerial biomass. In this study, the chemical composition of the root (crop-residue) essential oil was investigated for the first time, using column-chromatography, GC-FID, GC-MS, LC-QTOF, and NMR techniques, which led to the identification of twenty constituents, with isolation of (E)-2-(2',4'-hexadiynylidene)-1,6-dioxaspiro [4.5]dec-3-ene (D6). The D6 was evaluated in vitro for neuroinflammation and acetylcholinesterase inhibitory potential. It showed inhibition of neuroinflammation in a concentration-dependent manner with significant inhibition of pro-inflammatory cytokines (TNF-α and IL-6) in LPS-stimulated BV2 microglial cells. D6 did not have any significant effect on the viability of the cells at the therapeutic concentrations. D6 also has shown acetylcholinesterase inhibitory potential (51.90±1.19 %) at the concentration of log 106  nM. The results showed that D6 has a potential role in the resolution of neuroinflammation, and its acetylcholinesterase inhibitory potential directs further investigation of its role in the management of Alzheimer's disease-related pathogenesis.

Antibacterial Efficacy of ZnO/Bentonite (Clay) Nanocomposites against Multidrug-Resistant Escherichia coli.

The emergence of multidrug-resistant (MDR) bacteria has spurred the exploration of therapeutic nanomaterials such as ZnO nanoparticles. However, the inherent nonspecific toxicity of ZnO has posed a significant obstacle to their clinical utilization. In this research, we propose a novel approach to improve the selectivity of the toxicity of ZnO nanoparticles by impregnating them onto a less toxic clay mineral, Bentonite, resulting in ZB nanocomposites (ZB NCs). We hypothesize that these ZB NCs not only reduce toxicity toward both normal and carcinogenic cell lines but also retain the antibacterial properties of pure ZnO nanoparticles. To test this hypothesis, we synthesized ZB NCs by using a precipitation technique and confirmed their structural characteristics through X-ray diffraction and Raman spectroscopy. Electron microscopy revealed composite particles in the size range of 20-50 nm. The BET surface area of ZB NCs, within a relative pressure (P/P0) range of 0.407-0.985, was estimated to be 31.182 m2/g. Notably, 50 mg/mL ZB NCs demonstrated biocompatibility with HCT 116 and HEK 293 cell lines, supported by flow cytometry and fluorescence microscopy analysis. In vitro experiments further confirmed a remarkable five-log reduction in the population of MDR Escherichia coli in the presence of 50 mg/mL of ZB NCs. Antibacterial activity of the nanocomposites was also validated in the HEK293 and HCT 116 cell lines. These findings substantiate our hypothesis and underscore the effectiveness of ZB NCs against MDR E. coli while minimizing nonspecific toxicity toward healthy cells.

Mitochondrial atp1 mRNA knockdown by a custom-designed pentatricopeptide repeat protein alters ATP synthase.

Spontaneous mutations are rare in mitochondria and the lack of mitochondrial transformation methods has hindered genetic analyses. We show that a custom-designed RNA-binding pentatricopeptide repeat (PPR) protein binds and specifically induces cleavage of ATP synthase subunit1 (atp1) mRNA in mitochondria, significantly decreasing the abundance of the Atp1 protein and the assembled F1Fo ATP synthase in Arabidopsis (Arabidopsis thaliana). The transformed plants are characterized by delayed vegetative growth and reduced fertility. Five-fold depletion of Atp1 level was accompanied by a decrease in abundance of other ATP synthase subunits and lowered ATP synthesis rate of isolated mitochondria, but no change to mitochondrial electron transport chain complexes, adenylates, or energy charge in planta. Transcripts for amino acid transport and a variety of stress response processes were differentially expressed in lines containing the PPR protein, indicating changes to achieve cellular homeostasis when ATP synthase was highly depleted. Leaves of ATP synthase-depleted lines showed higher respiratory rates and elevated steady-state levels of numerous amino acids, most notably of the serine family. The results show the value of using custom-designed PPR proteins to influence the expression of specific mitochondrial transcripts to carry out reverse genetic studies on mitochondrial gene functions and the consequences of ATP synthase depletion on cellular functions in Arabidopsis.

Efficient and sustained FOXP3 locus editing in hematopoietic stem cells as a therapeutic approach for IPEX syndrome.

Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is a monogenic disorder caused by mutations in the FOXP3 gene, required for generation of regulatory T (Treg) cells. Loss of Treg cells leads to immune dysregulation characterized by multi-organ autoimmunity and early mortality. Hematopoietic stem cell (HSC) transplantation can be curative, but success is limited by autoimmune complications, donor availability and/or graft-vs.-host disease. Correction of FOXP3 in autologous HSC utilizing a homology-directed repair (HDR)-based platform may provide a safer alternative therapy. Here, we demonstrate efficient editing of FOXP3 utilizing co-delivery of Cas9 ribonucleoprotein complexes and adeno-associated viral vectors to achieve HDR rates of >40% in vitro using mobilized CD34+ cells from multiple donors. Using this approach to deliver either a GFP or a FOXP3 cDNA donor cassette, we demonstrate sustained bone marrow engraftment of approximately 10% of HDR-edited cells in immune-deficient recipient mice at 16 weeks post-transplant. Further, we show targeted integration of FOXP3 cDNA in CD34+ cells from an IPEX patient and expression of the introduced FOXP3 transcript in gene-edited primary T cells from both healthy individuals and IPEX patients. Our combined findings suggest that refinement of this approach is likely to provide future clinical benefit in IPEX.

Unveiling the cytotoxic and anti-proliferative potential of green-synthesized silver nanoparticles mediated by Colletotrichum gloeosporioides.

Fungal endophytes are a putative source of bioactive metabolites that have found significant applications in nanomedicine due to their metabolic versatility. In the present study, an aqueous extract of the fungal endophyte, Colletotrichum gloeosporioides associated with a medicinal plant Oroxylum indicum, has been used for the fabrication of green silver nanoparticles (CgAgNPs) and further evaluated their cytotoxic and anti-proliferative activity. Bioanalytical techniques including UV-Vis spectral analysis revealed a sharp band at 435 nm and functional molecules from the aqueous extract involved in the synthesis of CgAgNPs were evidenced through FTIR. Further, the crystalline nature of CgAgNPs was determined through XRD analysis and microscopy techniques including AFM, TEM and FESEM demonstrated the spherical shape of CgAgNPs exhibiting a crystalline hexagonal lattice and the size was found to be in the range of 9-29 nm. The significant cytotoxic potential of CgAgNPs was observed against breast cancer cells, MDA-MB-231 and MCF-7 with IC50 values of 18.398 ± 0.376 and 38.587 ± 1.828 µg mL-1, respectively. The biochemical study revealed that the treatment of MDA-MB-231 and MCF-7 cells with CgAgNPs reduces glucose uptake, suppresses cell proliferation, and enhances LDH release, indicating reduced cell viability and progression. Moreover, our research revealed differential expression of genes associated with apoptosis, cell cycle inhibition and metastasis suppression, evidencing anti-proliferative activity of CgAgNPs. The main objective of the present study is to harness anti-breast cancer activity of novel biogenic nanoparticles synthesized using the aqueous extract of O. indicum associated C. gloeosporioides and study the underlying mechanistic pathway exerted by these mycogenic nanoparticles.

Indian patients with CHST3-related chondrodysplasia with congenital joint dislocations.

CHST3-related chondrodysplasia with congenital joint dislocations (CDCJD, #MIM 143095), is a rare genetic skeletal disorder caused by biallelic loss of function variants in CHST3. CHST3 is critical for the sulfation of chondroitin sulfate. This study delineates the clinical presentation of nine individuals featuring the key symptoms of CDCJD; congenital joint (knee and elbow) dislocations, short trunk short stature progressive vertebral anomalies, and metacarpal shortening. Additional manifestations include irregular distal femoral epiphysis, supernumerary carpal ossification centers, bifid humerus, club foot, and cardiac abnormalities. Sanger sequencing was carried out to investigate molecular etiology in eight patients and exome sequencing in one. Genetic testing revealed five homozygous variants in CHST3 (four were novel and one was previously reported). All these variants are located on sulfotransferase domain of CHST3 protein and were classified as pathogenic/ likely pathogenic. We thus report on nine individuals with CHST3-related chondrodysplasia with congenital joint dislocations from India and suggest monitoring the health of cardiac valves in this condition.

Polyphenol modified CuO nanorods capped by kappa-carrageenan for controlled paclitaxel release in furnishing targeted chemotherapy in breast carcinoma cells.

The present work deals with the construction of a nanoscale system that can deliver chemotherapeutic agents to breast cancer cells in a controlled trend. The framework consists of gallic acid functionalized copper oxide nanoparticles (Ga@CuO) loaded with paclitaxel (PTX). To control the release of PTX, Ga@CuO NPs were coated with a red seaweed, Kappa carrageenan (K-carr) layer, and embellished with folic acid (FA) to enhance the targeted chemotherapy approach. Encapsulation efficiency and loading capacity of PTX loaded Ga@CuO@K-carr/FA NPs were estimated to be 84.58 ± 1.85 % and 13.2 ± 0.22 %, respectively. Moreover, the presence of strong cytotoxicity with an IC50 value of 12 ± 2.0 µg/mL and a high percentage of apoptotic cells (40.25 %) within the treated MCF-7 cells provided further evidence of the effective release of PTX from the loaded Ga@CuO@K-carr/FA. Consequently, it was discovered that the altered metabolic activity of cancer cells and the inhibition of cell proliferation are distinct features of apoptotic cell death induced by reactive oxygen species (ROS). Also, it was noted that treatment of MCF-7 cells with Ga@CuO-PTX@K-carr/FA caused a reduction in mitochondrial membrane potential that resulted in cellular apoptosis. Taken together, this study sheds light on the rational design of Ga@CuO-PTX@K-carr/FA, which offers a suitable candidate to transport drugs at an intracellular level for targeted chemotherapy.

New Insights Into the Anatomy of the Eyelashes With Regard to Ethnic Differences Against a Clinical Background.

BACKGROUND: To investigate the root depth, root angle, and light and scanning electron microscopical anatomy of human eyelashes relevant to eyelash ablation. METHODS: Eyelash root depth, the angle between eyelash root and skin epithelium, spatial relationship, and scanning electron microscopical features of the eyelashes were studied on 4 upper and 4 lower eyelids of Caucasian (n = 4) and Indian (n = 4) cadaver heads according to a set protocol. RESULTS: There were significant differences in the mean eyelash root depth between Indians (2.3 ± 0.38 mm) and Caucasians (1.9 ± 0.26 mm; p = 0.007), as well as between upper eyelids and lower eyelids (1.9 ± 0.2 mm vs. 1.8 ± 0.1 mm). The mean angle between the lash follicle root and the skin epithelium was 75 ± 11 degrees and similar in both ethnic groups. The eyelash bulb was located close to the tarsal plate and meibomian glands and formed an angle of less than 15 degrees with the eyelash root. Scanning electron microscopy studies revealed that the eyelash bulb was 202 ± 12 µm wide in Indians and 170.6 ± 16.8 µm wide in Caucasian eyelids ( p = 0.08). The eyelashes were placed more closely in Indian eyelids than in Caucasian eyelids ( p = 0.03). The width of the cuticle layer varied between the hair shaft and the inner eyelid segment. CONCLUSIONS: There are differences in eyelash root depth, inter-eyelash distance, and cuticle thickness between Indian and Caucasian eyelids. The oblique orientation of the eyelash root and close proximity of the eyelash bulb to the tarsal plate should be kept in mind while doing the electroepilation procedure.