Is There Any Association between Nutrition and Myopia? A Systematic Review.

SIGNIFICANCE: This systematic review highlights the possible role of nutrition in myopia based on qualitative analysis of vast and diverse literature that investigated this association. PURPOSE: We systematically reviewed the outcomes of the studies that previously investigated the association between nutrition and myopia. METHODS: EMBASE, MEDLINE, and PubMed were searched by two independent authors to identify cross-sectional, cohort, retrospective, or interventional studies that assessed the association of nutrition with myopia from inception to the year 2021. Furthermore, the reference list of the included articles was screened. The data from the included studies were extracted, and qualitative analysis was performed. Quality assessment for noninterventional studies and interventional trials was performed using the Newcastle-Ottawa Scale and Cochrane RoB 2, respectively. RESULTS: Twenty-seven articles were included in the review. Most of the nutrients and dietary elements investigated in noninterventional studies showed inconsistencies in their association with myopia, with the majority indicating no association. Nine studies showed a significant association of diverse nutrients and dietary elements with either an increase (odds ratio, 1.07) or a decrease (odds ratio, 0.5 to 0.96) in the risk of myopia development. However, a majority of these studies have minimal odds ratios with wider or overlapping confidence intervals, implicating weaker associations. All three nutrients and dietary elements assessed in the interventional trial had implications for myopia control, with two trials indicating a clinically minimal effect. CONCLUSIONS: This review implies that there is some evidence to indicate a potential influence of specific nutrients and dietary elements in myopia development, which are supported by several theories. However, given the vast, diverse, and complex nature of nutrition, more systematic investigation is warranted to comprehend the extent to which these specific nutrients and dietary elements are associated with myopia through longitudinal studies by subduing the limitations in the existing literature.

Myopia progression risk assessment score (MPRAS): a promising new tool for risk stratification.

Timely identification of individuals "at-risk" for myopia progression is the leading requisite for myopia practice as it aids in the decision of appropriate management. This study aimed to develop 'myopia progression risk assessment score' (MPRAS) based on multiple risk factors (10) to determine whether a myope is "at-risk" or "low-risk" for myopia progression. Two risk-score models (model-1: non-weightage, model-2: weightage) were developed. Ability of MPRAS to diagnose individual "at-risk" for myopia progression was compared against decision of five clinicians in 149 myopes, aged 6-29 years. Using model-1 (no-weightage), further 7 sub-models were created with varying number of risk factors in decreasing step-wise manner (1a: 10 factors to 1g: 4 factors). In random eye analysis for model-1, the highest Youden's J-index (0.63-0.65) led to the MPRAS cut-off score of 41.50-43.50 for 5 clinicians with a sensitivity ranging from 78 to 85% and specificity ranging from 79 to 87%. For this cut-off score, the mean area under the curve (AUC) between clinicians and the MPRAS model ranged from 0.89 to 0.90. Model-2 (weighted for few risk-factors) provided similar sensitivity, specificity, and AUC. Sub-model analysis revealed greater AUC with high sensitivity (89%) and specificity (94%) in model-1g that has 4 risk factors compared to other sub-models (1a-1f). All the MPRAS models showed good agreement with the clinician's decision in identifying individuals "at-risk" for myopia progression.

Zona pellucida glycoproteins: Relevance in fertility and development of contraceptive vaccines.

Mammalian zona pellucida (ZP) is composed of three to four glycoproteins, which plays an important role during fertilization. Mutations in the genes encoding zona proteins are reported in women with empty follicle syndrome, degenerated oocytes and those with an abnormal or no ZP further emphasizing their relevance during fertility. Immunization with either native or recombinant ZP glycoproteins/proteins leads to curtailment of fertility in various animal species. Observed infertility is frequently associated with ovarian pathology characterized by follicular atresia and degenerative changes in ZP, which may be due to oophoritogenic T cell epitope(s) within ZP glycoproteins. To avoid ovarian dystrophy, B cell epitopes of ZP glycoproteins have been mapped by using bio-effective monoclonal antibodies. Immunization with the immunogens encompassing the mapped B cell epitopes by and large led to amelioration of follicular atresia. However, their use for human application will require more rigorous research to establish their safety and reversibility of the contraceptive effect. Nonetheless, to minimize human-animal conflicts, ZP-based contraceptive vaccines have been used successfully in the population management of free-ranging animal species such as feral horses, white-tailed deer and elephants. To control zoonotic diseases, attempts are also underway to control the population of other animal species including stray dogs, which acts as one of the major vectors for the rabies virus.

Contraceptive efficacy of recombinant porcine zona proteins and fusion protein encompassing canine ZP3 fragment and GnRH in female beagle dogs.

PROBLEM: To manage population of dogs (Canis familiaris), the efficacy of recombinant proteins-based contraceptive vaccines to inhibit fertility has been evaluated in female beagle dogs. METHOD OF STUDY: Female beagle dogs (n = 4) were immunized with physical mixture of Escherichia coli-expressed recombinant porcine ZP3 with promiscuous T cell epitope of tetanus toxoid (TT-KK-pZP3) and porcine ZP4 with promiscuous T cell epitope of bovine RNase (bRNase-KK-pZP4), or with a fusion protein encompassing dog ZP3 fragment and two copies of GnRH with appropriate promiscuous T cell epitopes (dZP3-GnRH2 ); control animals received only alum, the adjuvant. The immunized animals were followed-up for antibody titres by ELISA as well as for fertility status subsequent to mating with male dogs. RESULTS: Active immunization of female dogs following a three injections schedule at 4-week intervals with a physical mixture of TT-KK-pZP3 + bRNase-KK-pZP4 as well as dZP3-GnRH2 , led to generation of significant antibody titres against respective recombinant proteins. Active immunization with dZP3-GnRH2 also led to generation of antibodies reactive with both dZP3 and GnRH. A booster dose on day 383 led to an increase in antibody titres and circulating antibodies against respective recombinant proteins could be observed on day 528. Antibodies in immune serum samples from dogs immunized with TT-KK-pZP3 + bRNase-KK-pZP4 or dZP3-GnRH2 reacted with native canine ZP as assessed by an indirect immunofluorescence assay. Mating studies revealed a reduced number of pregnancies as well as a significant reduction in the number of pups born in the female dogs immunized with dZP3-GnRH2 as compared to the adjuvanted control. Curtailment of pregnancy in dZP3-GnRH2 immunized group was associated with antibody titres against dZP3-GnRH2 . However, immunization with recombinant TT-KK-pZP3 + bRNase-KK-pZP4 did not significantly decrease the number of pups born as compared to the adjuvanted control. CONCLUSION: These studies revealed the potential of recombinant dZP3-GnRH2 -based contraceptive vaccine to curtail fertility in female dogs. Large scale studies to establish the efficacy and safety of this recombinant protein for the management of community dog population are thus warranted.

High stretchability, strength, and toughness of living cells enabled by hyperelastic vimentin intermediate filaments.

In many developmental and pathological processes, including cellular migration during normal development and invasion in cancer metastasis, cells are required to withstand severe deformations. The structural integrity of eukaryotic cells under small deformations has been known to depend on the cytoskeleton including actin filaments (F-actin), microtubules (MT), and intermediate filaments (IFs). However, it remains unclear how cells resist severe deformations since both F-actin and microtubules yield or disassemble under moderate strains. Using vimentin containing IFs (VIFs) as a model for studying the large family of IF proteins, we demonstrate that they dominate cytoplasmic mechanics and maintain cell viability at large deformations. Our results show that cytoskeletal VIFs form a stretchable, hyperelastic network in living cells. This network works synergistically with other cytoplasmic components, substantially enhancing the strength, stretchability, resilience, and toughness of cells. Moreover, we find the hyperelastic VIF network, together with other quickly recoverable cytoskeletal components, forms a mechanically robust structure which can mechanically recover after damage.

The Human Egg's Zona Pellucida.

Human zona pellucida (ZP) matrix, a delicate network of thin interconnected filaments, is primarily composed of four glycoproteins, namely, ZP1, ZP2, ZP3, and ZP4. All four zona proteins share common structural elements such as signal peptide, "ZP domain," consensus furin cleavage site, transmembrane-like domain, and short cytoplasmic tail. In addition, ZP1 and ZP4 also have "Trefoil domain." Recombinant/native human zona proteins have been used to investigate their binding characteristics to the capacitated and/or acrosome-reacted spermatozoa. These investigations revealed that ZP1, ZP3, and ZP4 primarily bind to the head region of the capacitated human spermatozoa, whereas ZP2 binds to the acrosome-reacted sperm. However, using transgenic mice, N-terminal region of human ZP2 has also been shown to play an important role in binding of sperm to the egg. ZP1, ZP3, and ZP4 lead to dose-dependent increase in acrosome reaction, suggesting that in humans more than one ZP glycoprotein is responsible for induction of acrosome reaction. Glycosylation of these proteins, in particular, N-linked glycosylation as well as sialyl-Lewisx, is essential for inducing acrosome reaction. Studies delineating downstream signaling events associated with induction of acrosome reaction reveal subtle differences between ZP3 and ZP1/ZP4 with respect to activation of Gi protein-coupled receptor and protein kinase A. The role of mutations in the zona proteins and ZP autoantibodies leading to infertility in women is suggestive and needs more rigorous experimentations for confirming their role in female infertility. The above-mentioned aspects of the human ZP glycoproteins have been discussed in this review.

Forskolin-mediated BeWo cell fusion involves down-regulation of miR-92a-1-5p that targets dysferlin and protein kinase cAMP-activated catalytic subunit alpha.

PROBLEM: To study the role of miRNA(s) during trophoblastic BeWo cell fusion. METHOD OF STUDY: Changes in miRNA(s) profile of BeWo cells treated with forskolin were analyzed using Affymetrix miRNA microarray platform. Down-regulated miRNA, miR-92a-1-5p, was overexpressed in BeWo cells followed by forskolin treatment to understand its relevance in the process of BeWo cell fusion by desmoplakin I+II staining and hCG secretion by ELISA. Predicted targets of miR-92a-1-5p were also confirmed by qRT-PCR/Western blotting. RESULTS: The miRNA profiling of BeWo cells after forskolin (25 µmol/L) treatment identified miR-92a-1-5p as the most significantly down-regulated miRNA both at 24 and 48 hours time points. Overexpression of miR-92a-1-5p in these cells led to a significant decrease in forskolin-mediated cell fusion and hCG secretion. miRNA target prediction software, TargetScan, revealed dysferlin (DYSF) and protein kinase cAMP-activated catalytic subunit alpha (PRKACA), as target genes of miR-92a-1-5p. Overexpression of miR-92a-1-5p in BeWo cells showed reduction in forskolin-induced transcripts for DYSF and PRKACA. Further, reduction in DYSF (~2.6-fold) at protein level and PRKACA-encoded protein kinase A catalytic subunit alpha (PKAC-α; ~1.6-fold) were also observed. CONCLUSION: These observations suggest that miR-92a-1-5p regulates forskolin-mediated BeWo cell fusion and hCG secretion by regulating PKA signaling pathway and dysferlin expression.

A simpler and more cost-effective peptide biosynthetic method using the truncated GST as carrier for epitope mapping.

There is a need to develop better methods for epitope mapping and/or identification of antibody-recognizing motifs. Here, we describe improved biosynthetic peptide (BSP) method using a newly developed plasmid pXXGST-3 as vector, which has a viral E7 gene in the cloning sites of pXXGST-1. It is crucial to employ pXXGST-3 instead of pXXGST-1, since it makes use of the BSP method simpler and easier to perform, and more cost-effective for epitope mapping. These merits are embodied in two aspects: i) convenient recovery of double enzyme-digested product due to the existence of 315 bp inserted between BamH I and Sal I sites, and thus greatly reducing the production of self-ligation clones, and ii) no longer requiring control protein when screening recombinant (r-) clones expressing 8/18mer peptides by running polyacrylamide gel electrophoresis. The protocol involves the following core steps: (i) design of plus and minus strands of DNA fragments encoding overlapping 8/18mer peptides; (ii) chemical synthesis of the designed DNA fragments; (iii) development of r-clones using pXXGST-3 vector expressing each 8/18mer peptide fused with truncated GST188 protein; (iv) screening r-clones by running the cell pellets from each induced clone on SDS-PAGE gel followed by sequencing of inserted DNA fragments for each verified r-clone; and (v) Western blotting with either monoclonal antibodies or polyclonal antibodies. This improved GST188-BSP method provides a powerful alternative tool for epitope mapping.

N-hydroxy-substituted 2-aryl acetamide analogs: A novel class of HIV-1 integrase inhibitors.

An in silico method has been used to discover N-hydroxy-substituted 2-aryl acetamide analogs as a new class of HIV-1 integrase inhibitors. Based on the molecular requirements of the binding pocket of catalytic active site, two molecules (compounds 2 and 4b) were designed as fragments. These were further synthesized and biologically evaluated. In vitro potency along with docking studies highlighted compound 4b as an active fragment which was further used to synthesize new leads as HIV-1 integrase inhibitors. Finally, six promising compounds (compounds 5b, 5c, 5e, 6-2c, 6-3b, and 6-5b) were identified by integrase inhibition assay (>50% inhibition). Based on in vitro anti-HIV-1 activity in a reporter gene-based cell assay system, compounds 5d, 6s, and 6k were found as novel HIV-1 integrase inhibitors due to its better selectivity index. Additionally, docking study revealed the importance of H-bond as well as hydrophobic interactions with Asn155, Lys156, and Lys159 which were required for their anti-HIV-1 activity.

Mapping Protein Binding Sites and Conformational Epitopes Using Cysteine Labeling and Yeast Surface Display.

We describe a facile method for mapping protein:ligand binding sites and conformational epitopes. The method uses a combination of Cys scanning mutagenesis, chemical labeling, and yeast surface display. While Ala scanning is widely used for similar purposes, often mutation to Ala (or other amino acids) has little effect on binding, except at hotspot residues. Many residues in physical contact with a binding partner are insensitive to substitution with Ala. In contrast, we show that labeling of Cys residues in a binding site consistently abrogates binding. We couple this methodology to yeast surface display and deep sequencing to map conformational epitopes targeted by both monoclonal antibodies and polyclonal sera as well as a protein:ligand binding site. The method does not require purified protein, can distinguish buried and exposed residues, and can be extended to other display formats, including mammalian cells and viruses, emphasizing its wide applicability.

Epitomics: IgG-epitome decoding of E6, E7 and L1 proteins from oncogenic human papillomavirus type 58.

To enable rational multi-epitope vaccine and diagnostic antigen design, it is imperative to delineate complete IgG-epitome of the protein. Here, we describe results of IgG-epitome decoding of three proteins from high-risk (HR-) oncogenic human papillomavirus type 58 (HPV58). To reveal their entire epitomes, employing peptide biosynthetic approach, 30 precise linear B-cell epitopes (BCEs) were mapped on E6, E7 and L1 proteins using rabbits antisera to the respective recombinant proteins. Using sequence alignment based on BCE minimal motif, the specificity and conservativeness of each mapped BCE were delineated mainly among known HR-HPVs, including finding 3 broadly antibody cross-reactive BCEs of L1 that each covers almost all HR-HPVs. Western blots revealed that 13 of the 18 BCEs within L1-epitome were recognized by murine antisera to HPV58 virus-like particles, suggesting that these are antibody accessible BCEs. Also, a highly conserved epitope (YGD/XTL) of E6 was found to exist only in known common HR-HPVs, which could be used as the first peptide reference marker for judging HR-HPVs. Altogether, this study provides systemic and exhaustive information on linear BCEs of HR-HPV58 that will facilitate development of novel multi-epitope diagnostic reagents/chips for testing viral antibodies and 'universal' preventive HPV peptide vaccine based on L1 conserved BCEs.

Efficiency of the occlusion therapy with and without levodopa-carbidopa in amblyopic children-A tertiary care centre experience.

OBJECTIVES: To assess the role, efficacy and tolerability of levodopa-carbidopa in the management of small and older children with different types of amblyopia. METHODOLOGY: Prospective randomised placebo controlled clinical study, in which 50 amblyopic patients between 5 and 20 years of age with visual acuity (V/A) < 20/40 were included, was carried on. After having attained the best possible refractive correction, patients were randomly divided into 2 groups. They were prescribed levodopa-carbidopa (10:1) (4-6mg/kg/day in 2-3 divided doses) or placebo, plus full-time occlusion of the sound eye, for a period of three months. Assessment of improvement in V/A, compliance and tolerance was done at follow up visits. Data was analyzed using computer software Ms-Excel and Epi-Info Version 6.0. The statistical significance was assessed by Chi-Square/Fisher's Exact Test. RESULTS: Visual acuity for the amblyopic eye improved significantly in both groups but there was significant improvement in group 1 than group 2 (P = 0.0001). In a subgroup of patients older than 12 years, levodopa group showed statistically significant improvement in baseline V/A (P = 0.0001). In patients with severe amblyopia, each group showed significant improvement in baseline V/A (p < 0.05), but was significantly more in group 1 (P = 0.0001). Compliance rates were similar among the groups and levodopa-carbidopa at a dose range of 4-6 mg/kg/day was well tolerated. CONCLUSION: Levodopa-carbidopa can be used as an adjunct to conventional occlusion therapy in amblyopia particularly in older children and severe cases of amblyopia, and it is well tolerated.

Synthesis, biological evaluation and molecular modeling studies of new 2,3-diheteroaryl thiazolidin-4-ones as NNRTIs.

In a focused exploration, thiazolidin-4-ones with different C-2 and N-3 substituent groups were synthesized and evaluated as non-nucleoside reverse transcriptase inhibitors against HIV-1. This has led to new active compounds sporting heteroaryls at both C-2 and N-3 positions prompting to view them in the backdrop of nevirapine. To assign the molecular attributes for the activity, the compounds are investigated by docking them into non-nucleoside inhibitor-binding pocket of HIV-1 reverse transcriptase (RT). The most active compounds of this series (7d and 7f) shared spatial features with nevirapine with added molecular flexibility. Furthermore, in molecular dynamics simulations carried out for up to 10 ns, the compounds 7d and 7f showed consistency in their interactions with non-nucleoside inhibitor-binding pocket of HIV-1 RT and suggested Tyr319 and Val106 as potential residues for H-bond interaction with these molecules. These results open new avenues for the exploration of 2,3-diheteroaryl thiazolidin-4-ones for prevention of HIV-1.

Unraveling the intricacies of mammalian fertilization.

It is imperative to understand the molecular basis of various steps involved during fertilization. In the manuscript by Bianchi et al. a novel protein, Juno on egg membrane (oolemma) has been characterized that binds to sperm specific protein, Izumo-1. Monoclonal antibodies against Juno inhibited in vitro fertilization. Juno knock-out female mice failed to deliver litters on mating. It is rapidly shed from oolemma after fertilization, suggesting its role in preventing polyspermy. Taken together these studies will help in our understanding of sperm-egg recognition mechanisms and also facilitate development of new fertility treatment regimens and novel contraceptives.

Synthesis, anti-HIV activity, integrase enzyme inhibition and molecular modeling of catechol, hydroquinone and quinol labdane analogs.

Labdane analogs with o-quinol, catechol and hydroquinone moiety have been synthesized using Diels-Alder reaction of methyl 3,4-dioxocyclohexa-1,5-diene-carboxylate, 3,4-dioxocyclohexa-1,5-diene-carboxylic acid and 3,6-dioxocyclohexa-1,4-dienecarboxylic acid with mono terpene 1,3-dienes, namely ocimene and myrcene. The resulting molecules and their derivatives were evaluated for their anti-HIV-1 activity using TZM-bl cell based virus infectivity assay. Two molecules 13 and 18 showed anti-HIV activity with IC50 values 5.0 (TI=11) and 4.6 (TI=46)µM, respectively. The compounds 17, 18 and 20 showed efficacy against HIV-1 integrase activity and showed inhibition with IC50 13.4, 11.1 and 11.5µM, respectively. The HIV-1 integrase inhibition activity of these synthetic molecules was comparable with integric acid, the natural fungal metabolite. Molecular modeling studies for the HIV-1 integrase inhibition of these active synthetic molecules indicated the binding to the active site residues of the enzyme.

Immunogenicity and contraceptive efficacy of Escherichia coli-expressed recombinant porcine zona pellucida proteins.

PROBLEM: To overcome availability of the purified native zona pellucida (ZP) glycoproteins for immunocontraception, porcine ZP3, and ZP4 were expressed in E. coli. METHOD OF STUDY: Purified recombinant proteins were characterized by SDS-PAGE and Western blot, and immunogenicity and contraceptive efficacy determined in FvB/J female mice. RESULTS: Purified ZP3, ZP3 with promiscuous T-cell epitope of tetanus toxoid, ZP4 and ZP4 incorporating promiscuous T-cell epitope of bovine RNase revealed ~44-, ~49-, ~53-, and ~55-kDa bands by SDS-PAGE and Western blot, respectively. Immunization of female mice with recombinant proteins elicited high antibody titers as well as T-cell responses. Immune sera recognized mouse oocyte ZP and also inhibited in vitro fertilization. Immunized mice showed significant decrease in fertility. Recombinant proteins were able to recall memory antibody response in female mice primed with porcine native ZP. CONCLUSION: Availability of recombinant porcine proteins will be useful in the development of contraceptive vaccine.

Pharmaceutical solid dispersion technology: a strategy to improve dissolution of poorly water-soluble drugs.

Oral bioavailability is the major problem when a poorly water-soluble active agent is delivered via oral route. To overcome such problems, solid dispersion systems have been demonstrated in literature to enhance the dissolution property of poorly water-soluble drugs. In the present review, the important aspects to be considered during preparation of solid dispersion systems viz., properties of polymer and preparation techniques of solid dispersion which affect the dissolution rate are discussed. Formulation and evaluation techniques for solid dispersions have been described. The final section of article highlights the recent patents and studies related to solid dispersion systems.

Haemophilus influenza type b disease and vaccination in India: knowledge, attitude and practices of paediatricians.

BACKGROUND: Haemophilus influenza type b (Hib) causes significant morbidity and mortality among young children in India. Hib vaccines are safe and efficacious; nevertheless, their introduction to India's national immunization programme has been hindered by resistance from certain sectors of academia and civil society. We aimed to ascertain the attitudes and perceptions of Indian paediatricians towards Hib disease and vaccination. MATERIALS AND METHODS: A cross-sectional survey of knowledge, attitude and practices on Hib and vaccines was undertaken among 1000 Indian paediatricians who attended 49 th National Conference of Indian Academy of Pediatrics in 2012 through use of a 21-point questionnaire. RESULTS: 927 (93%) paediatricians completed the survey. 643 (69%) responded that Hib is a common disease in India. 788 (85%) reported prescribing Hib vaccine to their patients and 453 (49%) had done so for the past 5-15 years. Hib vaccine was used in combination with other vaccines by 814 (88%) of the participants. 764 (82%) respondents thought Hib vaccine effective while 750 (81%) thought it to be safe. Fever, pain and redness were the most frequently reported post vaccination side-effects. 445 (48%) paediatricians ranked universal use of Hib vaccine in the national immunization programme as the most important strategy to prevent and control Hib disease in India. CONCLUSION: The excellent profile as reported by a large number of paediatricians from throughout India further strengthens evidence to support expanded use of currently available Hib vaccines. These findings should encourage the Government of India to initiate mass use of this vaccine nationwide.

Mapping of epitopes relevant for induction of acrosome reaction on human zona pellucida glycoprotein-4 using monoclonal antibodies.

PROBLEM: To decipher structural and functional aspects of human zona pellucida glycoprotein-4 (ZP4), the epitopes recognized by monoclonal antibodies (MAbs) have been mapped. METHOD OF STUDY: Recombinant human ZP4-mediated induction of acrosome reaction in human sperm was studied in the absence and presence of ZP4-specific MAbs. The epitopes of MAbs were mapped using recombinant peptides expressed in Escherichia coli. RESULTS: Monoclonal antibodies (MA-1662, MA-1671) against human ZP4 showed specific binding to ZP matrix of human eggs in an indirect immunofluorescence assay. Both the antibodies showed significant (P < 0.05) inhibition in the baculovirus-expressed recombinant ZP4-mediated acrosome reaction. MA-1671 recognized N-terminal fragment of ZP4 and minimal epitope mapped to amino acid residues 126-130 (PARDR), whereas MA-1662 reacted to C-terminal fragment and minimal epitope mapped to amino acid residues 256-260 (ENELV). CONCLUSIONS: The epitopes corresponding to both N- and C-terminal parts of human ZP4 may be relevant for its biological activity.