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An experiment using cv. Taiwan was 786 laid out in factorial completely randomized design with thirty treatments and three replications was, carried out at Laboratory of Horticulture Section, College of Agriculture, Dhule (M.S.) during 2017. The experiment comprised of three factors viz. two maturity stages (25% mature and 50% mature), dipping them in aqueous solutions of spermine (1.0 mM and 2.0 mM) and calcium lactate (1% and 2%) for 5 minutes and drying for 30 minutes at room temperature, three packaging materials (newspaper, shrink film and polyethylene bags) were used. Results indicated a significant impact of interaction among maturity stages, spermine and calcium lactate and packaging material on all parameters included in the study. When considered interaction, twenty five percent mature fruits dipped in (2.0 mM) spermine with shrink packaging had the minimum physiological loss in weight. This treatment also recorded the maximum peel to pulp ratio, fruit firmness. The maximum shelf life of 16 days without sacrificing organoleptic quality by manipulating the time of harvesting, packaging and application of spermine. [ABSTRACT FROM AUTHOR]

Objectives: Among infectious diseases, bloodstream infection (BSI) caused by gram-negative bacteria (E. coli) is the leading cause of death worldwide. However, the bacteria have produced resistance to many of these antibiotics. Thus, the present study aimed to develop silver nanoparticles (AgNPs) loaded with Emilia sonchifolia (ES) extract (ES-AgNPs) to treat BSI efficiently., Methods: AgNPs were synthesized by reduction of silver nitrate (AgNO 3 ) solution by ES extract. Furthermore, these ES-AgNPs were characterized for particle size and zeta potential, crystallinity by powder X-ray diffraction (P-XRD) technique, in vitro antibacterial activity, time-kill assay, film bio adhesion, and fluorescence assay., Results: Surface plasmon resonance (SPR) has been used to confirm the formation of AgNPs by seeing a shift in colour to dark-brown. The ES-AgNPs displayed a mean particle size of 137±3nm (PDI of 0.168±0.02) and zeta potential of 18.2±0.8mV. Furthermore, according to P-XRD results, the developed AgNPs are highly crystalline. The ES-AgNPs showed effective antibacterial action against E. coli with minimum inhibitory concentrations (MIC) and minimum bactericidal concentrations (MBC) of 0.4±0.02μg/mL and 0.8±0.03μg/mL, respectively. In addition, ES-AgNPs inhibited biofilm formation and bacterial adhesion in a dose-dependent manner with 100% inhibition obtained in 48h at MBC., Conclusions: Present research work revealed that the ES-AgNPs obtained by green synthesis holds a prominent antibacterial activity in the treatment of BSIs caused by E. coli and they may be used as a competent substitute for current treatments. However, further, in vivo antibacterial studies are required to establish its efficacy in the treatment of BSIs., (Copyright © 2022 Académie Nationale de Pharmacie. Published by Elsevier Masson SAS. All rights reserved.)

Purpose: The outflow facility for aqueous humor across the trabecular meshwork (TM) is enhanced by agents that oppose the actomyosin contraction of its resident cells. Phosphorylation of MYPT1 (myosin light chain [MLC] phosphatase complex of Type 1) at Thr853 and Thr696 inhibits dephosphorylation of MLC, leading to an increase in actomyosin contraction. In this study, we examined the effects of Rho kinase (ROCK) inhibitors on the relative dephosphorylation of the two sites of MYPT1 using human TM cells (GTM3)., Methods: Dephosphorylation of MYPT1 at Thr853 and Thr696 was determined by western blot analysis following exposure to selective inhibitors of ROCK, namely Y-27632 and Y-39983. Consequent dephosphorylation of MLC and decreases in actomyosin contraction were assessed by western blot analysis and collagen gel contraction assay, respectively. Changes in the cell-matrix adhesion were measured in real time by electric cell-substrate impedance sensing and also assessed by staining for paxillin, vinculin, and focal adhesion kinase (FAK)., Results: Both ROCK inhibitors produced a concentration-dependent dephosphorylation at Thr853 and Thr696 of MYPT1 in adherent GTM3 cells. IC₅₀ values for Y-39983 were 15 nM and 177 nM for dephosphorylation at Thr853 and Thr696, respectively. Corresponding values for Y-27632 were 658 nM and 2270 nM. Analysis of the same samples showed a decrease in MLC phosphorylation with IC₅₀ values of 14 nM and 1065 nM for Y-39983 and Y-27632, respectively. Consistent with these changes, both inhibitors opposed contraction of collagen gels induced by TM cells. Exposure of cells to the inhibitors led to a decrease in the electrical cell-substrate resistance, with the effect of Y-39983 being more pronounced than Y-27632. Treatment with these ROCK inhibitors also showed a loss of stress fibers and a concomitant decrease in tyrosine phosphorylation of paxillin and FAK., Conclusions: Y-39983 and Y-27632 oppose ROCK-dependent phosphorylation of MYPT1 predominantly at Thr853 with a corresponding decrease in MLC phosphorylation. A relatively low effect of both ROCK inhibitors at Thr696 suggests a role for other Ser/Thr kinases at this site. Y-39983 was several-fold more potent when compared with Y-27632 at inhibiting the phosphorylation of MYPT1 at either Thr853 or Thr696 commensurate with its greater potency at inhibiting the activity of human ROCK-I and ROCK-II enzymes.

We report for the first time that cultured nonpigmented human ciliary epithelial (NPE) cell layers transport fluid. Cells were grown to confluence on permeable membrane inserts, and fluid transport across the resulting cell layers was determined by volume clamp at 37 degrees C. These cell layers translocated fluid from the apical to the basal side at a steady rate of 3.6 microl x h(-1) x cm(-2) (n = 4) for 8 h. This fluid movement was independent of hydrostatic pressure and was completely inhibited by 1 mM ouabain, suggesting it arose from fluid transport. Mercuric chloride, a nonspecific but potent blocker of Hg(2+)-sensitive aquaporins, and aquaporin-1 antisense oligonucleotides both partially inhibited fluid transport across the cell layers, which suggests that water channels have a role in NPE cell homeostasis. In addition, these results suggest that of the two ciliary epithelial layers in tandem, the NPE layer by itself can transport fluid. This cultured layer, therefore, constitutes an interesting model that may be useful for physiological and pharmacological characterization of ciliary epithelial fluid secretion.

Purpose: To determine the expression and localization of tumor necrosis factor (TNF)-alpha and TNF-alpha receptor-1 in the retina of normal and glaucomatous eyes., Methods: Using immunohistochemistry and in situ hybridization, retinal expression and localization of TNF-alpha and TNF-alpha receptor-1 were studied in retina sections from 20 eyes of donors with glaucoma, and 20 eyes of age-matched normal donors., Results: According to immunohistochemistry, the intensity of the immunostaining and the number of labeled cells for TNF-alpha or its receptor were greater in retina sections of glaucomatous eyes than in control eyes of age-matched normal donors. In situ hybridization showed that mRNA signals for TNF-alpha or TNF-alpha receptor-1 were similarly more intense in glaucomatous eyes than in age-matched control eyes. Both protein and mRNA of TNF-alpha or TNF-alpha receptor-1 were predominantly localized to the inner retinal layers. Double-immunofluorescence labeling demonstrated that retinal immunostaining for TNF-alpha was predominantly positive in the glial cells, whereas immunostaining for TNF-alpha receptor-1 was mainly positive in the retinal ganglion cells., Conclusions: Upregulation of TNF-alpha and its receptor-1 in glaucomatous retina suggest that TNF-alpha-mediated cell death is involved in the neurodegeneration process of glaucoma.

Purpose: To identify retinal proteins that are the targets of serum autoantibodies in patients with glaucoma., Methods: To identify retinal antigens that are recognized by the sera of patients with glaucoma, immunoreactive bands were separated, by using two-dimensional gel electrophoresis of the bovine retinal soluble fraction. A 29-kDa band was then selected for further analysis. Tryptic peptides of the 29-kDa band were analyzed using electrospray mass spectrometry to identify the protein. After protein identification, immunoreactivity against this newly identified protein was studied by Western blot analysis using sera from 65 patients with glaucoma (25 with primary open-angle glaucoma [POAG]; 40 with normal-pressure glaucoma [NPG]) and 25 age-matched healthy subjects. In addition, serum antibody titers were compared in these groups, by using a specific enzyme-linked immunosorbent assay (ELISA)., Results: The 29-kDa band was identified as glutathione S-transferase (GST). Western blot analysis revealed that serum antibodies against GST antigen were recognized in 34 (52%) of 65 patients with glaucoma (22 of NPG and 12 of POAG) and 5 (20%) of 25 age-matched control subjects (chi(2) test, P < 0.05). By ELISA, it was also found that patients with glaucoma had higher titers of anti-GST antibody, compared with the control group (Mann-Whitney test; NPG versus control, P = 0.013; POAG versus control, P = 0.0006)., Conclusions: These findings indicate that GST is one of the retinal antigens targeted by the serum antibodies detected in some patients with glaucoma.

Purpose: Recent evidence strongly suggests that activated immunity occurs during the neurodegenerative process of glaucomatous optic neuropathy. Although activation of lamina cribrosa astrocytes has been identified in glaucomatous optic nerve head, their role on the activated immune responses seen in glaucoma patients is unknown. Here, the authors aimed to study the potential role of lamina cribrosa astrocytes as a component of activated immune responses seen in glaucoma patients., Methods: Expression of HLA-DR in optic nerve head astrocytes was studied using immunohistochemistry in postmortem eyes of patients with glaucoma and normal donors. Serum cytokine levels of patients with glaucoma and control subjects were measured using enzyme-linked, immunosorbent assay. In addition, in vitro experiments were performed using astrocyte cultures derived from human optic nerve head or fetal human brain. The cultured astrocytes were incubated under selected stress conditions such as exposure to cytokines, IFN-gamma and IL-10, or simulated ischemia for up to 48 hours. The expression of HLA-DR was studied in these cells using flow cytometry and immunocytochemistry., Results: Immunohistochemistry demonstrated an upregulation of the HLA-DR expression in the optic nerve head astrocytes in glaucoma. In addition, serum levels of IL-10 was higher in the patients with normal pressure glaucoma compared to age-matched control subjects (P: = 0.001). Regarding in vitro experiments, unlike brain astrocytes, the percentage of cells expressing HLA-DR was approximately 3 times higher in the cultures of optic nerve head astrocytes exposed to simulated ischemia compared to cultures incubated under normal conditions (P: = 0.09). Incubation with IFN-gamma induced HLA-DR expression in brain and lamina cribrosa astrocytes, up to 25-fold, (P < 0.001) either in the absence or presence of simulated ischemia. Induction of HLA-DR expression by IL-10 was approximately 6 times higher in lamina cribrosa astrocytes incubated under simulated ischemia compared to that incubated under normal condition (P: = 0.004) and was not prominent in brain astrocytes., Conclusions: These findings suggest that optic nerve head astrocytes function as antigen-presenting cells and that their immunogenic capacity is more sensitive to ischemia than brain astrocytes. Taken together, these findings provide novel evidence that regulation of immunogenic capacity of optic nerve head astrocytes by cytokines or ischemic stress may have a role during the neurodegeneration process in patients with glaucoma.

Purpose: To count retrogradely labeled retinal ganglion cells by Fluoro-Gold., Methods: Retinal ganglion cells were retrogradely labeled using bilateral injections of Fluoro-Gold into the superior colliculus. One week after injections, retinas were dissociated and immunolabeled using specific antibody against Fluoro-Gold. The Fluoro-Gold labeled cells were then counted using flow cytometry., Results: Flow cytometry revealed that approximately 7% of the dissociated retinal cells were ganglion cells retrogradely labeled by Fluoro-Gold (Fig. 1B). Based on the total count of retinal cells per eye, the total number of retinal ganglion cells was estimated at approximately 131,250 +/- 2,542 per rat eye. The coefficient of variation of counts was calculated as 1.98%., Conclusions: The use of flow cytometry facilitates simple, reproducible and rapid quantification of virtually all of the retinal ganglion cells labeled by Fluoro-Gold in a single rat eye.