Multimerin 1 aids in the progression of ovarian cancer possibly via modulation of DNA damage response and repair pathways.

Ovarian cancer is one of the leading causes of deaths among women. Despite advances in the treatment regimes, a high rate of diagnosis in the advanced stage makes it almost an incurable malignancy. Thus, more research efforts are required to identify potential molecular markers for early detection of the disease and therapeutic targets to augment the survival rate of ovarian cancer patients. Previously, in this context, we identified dysregulated expression of multimerin 1 (MMRN1) in ovarian cancer. To elucidate the relationship between MMRN1 expression and ovarian cancer progression, siRNA-based MMRN1 knockdown was employed and various cell assays were performed to study its effect on ovarian cancer cells. In addition, network of dysregulated proteins was identified by quantitative proteomics and associated pathways were explored by bioinformatics analysis. MMRN1 silencing showed a significant reduction in cell viability, adhesion, migration, and invasion and a high frequency of cell apoptosis. Label-free quantitative proteomics and in-depth statistical analysis identified 448 dysregulated proteins, majority of which were overexpressed in MMRN1 knockdown cells. The pathways overrepresented in ovarian cancer were DNA replication, mismatch repair, nucleotide excision repair, and cell cycle regulation. Conclusively, the findings of this study suggest that MMRN1 aids in the progression of ovarian cancer via modulation of DNA damage response and repair pathways.

Unveiling the Significance of FGF8 Overexpression in Orchestrating the Progression of Ovarian Cancer.

The asymptomatic nature, high rate of disease recurrence, and resistance to platinum-based chemotherapy highlight the need to identify and characterize novel target molecules for ovarian cancer. Fibroblast growth factor 8 (FGF8) aids in the development and metastasis of ovarian cancer; however, its definite role is not clear. We employed ELISA and IHC to examine the expression of FGF8 in the saliva and tissue samples of epithelial ovarian cancer (EOC) patients and controls. Furthermore, various cell assays were conducted to determine how FGF8 silencing influences ovarian cancer cell survival, adhesion, migration, and invasion to learn more about the functions of FGF8. In saliva samples, from controls through low-grade to high-grade EOC, a stepped overexpression of FGF8 was observed. Similar expression trends were seen in tissue samples, both at protein and mRNA levels. FGF8 gene silencing in SKOV3 cells adversely affected various cell properties essential for cancer cell survival and metastasis. A substantial reduction was observed in the cell survival, cell adhesion to the extracellular matrix, migration, and adhesion properties of SKOV3 cells, suggesting that FGF8 plays a crucial role in the development of EOC. Conclusively, this study suggests a pro-metastatic function of FGF8 in EOC.

Analysis of Multimerin 1 (MMRN1) expression in ovarian cancer.

Ovarian cancer, the most lethal gynecological cancer, is the fifth most common cause of cancer-related deaths in women. A cost-effective and non-invasive early screening method for ovarian cancer is required to reduce the high mortality rate. Saliva is a clinically informative unique fluid, which is useful for novel approaches to prognosis, clinical diagnosis, and monitoring for non-invasive detection of disease. Multimerin1 (MMRN1) is a di-sulfide linked homo-polymeric glycoprotein from EMILIN family. Altered expression of MMRN1 has been reported in hepatocellular carcinoma, cervical cancer, and ovarian cancer. But, its role in epithelial ovarian cancer (EOC) is not clear and well documented. In this study, expression of Multimerin 1 was validated in saliva and tissues of EOC patients and age-matched controls by western blotting, ELISA, RT-PCR, and immunohistochemistry. Significant over expression of MMRN1 was observed by western blot and ELISA in saliva samples of EOC patients. The average concentration of MMRN1 in the saliva of healthy controls was 28.7 pg/ml (SE ± 1.76), 42.53 pg/ml (SE ± 4.06) in low grade and 52.91 pg/ml (SE ± 4.24) with p < 0.01 in high-grade EOC. Upregulated cytoplasmic expression of MMRN1 was observed in EOC tissue by immunohistochemistry. Our results suggest that MMRN1 expression is associated with EOC progression and MMRN1 may be potential biomarker candidates for early-stage EOC detection however further experiments are required in a large cohort to establish this proposition. Also, saliva can be explored as a novel medium for ovarian cancer diagnosis.

Human Spermatozoa Quantitative Proteomic Signature Classifies Normo- and Asthenozoospermia.

Scarcely understood defects lead to asthenozoospermia, which results in poor fertility outcomes. Incomplete knowledge of these defects hinders the development of new therapies and reliance on interventional therapies, such as in vitro fertilization, increases. Sperm cells, being transcriptionally and translationally silent, necessitate the proteomic approach to study the sperm function. We have performed a differential proteomics analysis of human sperm and seminal plasma and identified and quantified 667 proteins in sperm and 429 proteins in seminal plasma data set, which were used for further analysis. Statistical and mathematical analysis combined with pathway analysis and self-organizing maps clustering and correlation was performed on the data set.It was found that sperm proteomic signature combined with statistical analysis as opposed to the seminal plasma proteomic signature can differentiate the normozoospermic versus the asthenozoospermic sperm samples. This is despite the results that some of the seminal plasma proteins have big fold changes among classes but they fall short of statistical significance. S-Plot of the sperm proteomic data set generated some high confidence targets, which might be implicated in sperm motility pathways. These proteins also had the area under the curve value of 0.9 or 1 in ROC curve analysis.Various pathways were either enriched in these proteomic data sets by pathway analysis or they were searched by their constituent proteins. Some of these pathways were axoneme activation and focal adhesion assembly, glycolysis, gluconeogenesis, cellular response to stress and nucleosome assembly among others. The mass spectrometric data is available via ProteomeXchange with identifier PXD004098.

Comparative proteomic analysis of high- and low-fertile buffalo bull spermatozoa for identification of fertility-associated proteins.

The present study identified few potential proteins in the spermatozoa of buffalo bulls that can be used as an aid in fertility determination through comparative proteomics. The sperm proteome of high-fertile buffalo bulls was compared with that of low-fertile buffalo bulls using two-dimensional difference gel electrophoresis (2D-DIGE), and the differentially expressed proteins were identified through mass spectrometric method. The protein interaction network and the functional bioinformatics analysis of differentially expressed proteins were also carried out. In the spermatozoa of high-fertile bulls, 10 proteins were found overexpressed and 15 proteins were underexpressed at the level of twofold or more (p ≤ 0.05). The proteins overexpressed in high-fertile spermatozoa were PDZD8, GTF2F2, ZNF397, KIZ, LOH12CR1, ACRBP, PRSS37, CYP11B2, F13A1 and SPO11, whereas those overexpressed in low-fertile spermatozoa were MT1A, ATP5F1, CS, TCRB, PRODH2, HARS, IDH3A, SRPK3, Uncharacterized protein C9orf9 homolog isoform X4, TUBB2B, GPR4, PMP2, CTSL1, TPPP2 and EGFL6. The differential expression ranged from 2.0- to 6.1-fold between the two groups, where CYP11B2 was high abundant in high-fertile spermatozoa and MT1A was highly abundant in low-fertile spermatozoa. Most of the proteins overexpressed in low-fertile spermatozoa were related to energy metabolism and capacitation factors, pointing out the possible role of pre-mature capacitation and cryo-damages in reducing the fertility of cryopreserved buffalo spermatozoa.

Efficacy and Safety of a Novel Method of Insulated Intralesional Radiofrequency Ablation for Deep Dermal and Subcutaneous Lesions: A 3-Year Institutional Experience.

BACKGROUND: Although insulated intralesional radiofrequency ablation (IL-RFA) is being increasingly used in other specialties, not much information on its safety and efficacy in dermatology is available. OBJECTIVE: To describe our experience with insulated IL-RFA for various dermatological conditions. METHODS: This is a retrospective review of the patients who underwent IL-RFA in the past 3 years. Our technique involved creating a small window in the proximal end of plastic sheath of an intravenous cannula using a surgical blade, and then touching the RF probe to the cannula through the window to deliver the electric current. Information regarding diagnosis, number of sessions, adverse effects, and follow-up was recorded. Clinical improvement was assessed on a visual analog scale by the patient and 2 independent observers. RESULTS: Data on 19 patients with lymphangioma circumscriptum (n = 9), venous or capillary-venous malformation (n = 4), angiolymphoid hyperplasia with eosinophilia (n = 3), arteriovenous malformation, hidradenitis suppurativa, and hypertrophic scar (n = 1 each) was available. The mean number of IL-RFA sessions was 2.26 ± 1.61. The mean patient and physician global assessment scores were 7.6 ± 2.22 and 7.3 ± 2.42, respectively. Adverse effects were seen in 9 (47.4%) patients. All patients, except 1, had sustained improvement in the mean follow-up period of 11.4 ± 11.6 months. CONCLUSION: Insulated IL-RFA seems to be safe and effective in selectively targeting deep-seated cutaneous lesions.

Children aged 6-24 months like to watch YouTube videos but could not learn anything from them.

AIM: Parents sometimes show young children YouTube videos on their smartphones. We studied the interaction of 55 Indian children born between December 2014 and May 2015 who watched YouTube videos when they were 6-24 months old. METHODS: The children were recruited by the researchers using professional and personal contacts and visited by the same two observers at four ages, for at least 10 minutes. The observers recorded the children's abilities to interact with touch screens and identify people in videos and noted what videos attracted them the most. RESULTS: The children were attracted to music at six months of age and were interested in watching the videos at 12 months. They could identify their parents in videos at 12 months and themselves by 24 months. They started touching the screen at 18 months and could press the buttons that appeared on the screen, but did not understand their use. The children preferred watching dance performances by multiple artists with melodical music, advertisements for products they used and videos showing toys and balloons. CONCLUSION: Children up to two years of age could be entertained and kept busy by showing them YouTube clips on smartphones, but did not learn anything from the videos.

Structural insights of Mycobacterium GTPase-Obg and anti-sigma-F factor Usfx interaction.

An essential protein for bacterial growth, GTPase-Obg (Obg), is known to play an unknown but crucial role in stress response as its expression increases in Mycobacterium under stress conditions. It is well reported that Obg interacts with anti-sigma-F factor Usfx; however, a detailed analysis and structural characterization of their physical interaction remain undone. In view of above-mentioned points, this study was conceptualized for performing binding analysis and structural characterization of Obg-Usfx interaction. The binding studies were performed by surface plasmon resonance, while in silico docking analysis was done to identify crucial residues responsible for Obg-Usfx interaction. Surface plasmon resonance results clearly suggest that N-terminal and G domains of Obg mainly contribute to Usfx binding. Also, binding constants display strong affinity that was further evident by intermolecular hydrogen bonds and hydrophobic interactions in the predicted complex. Strong interaction between Obg and Usfx supports the view that Obg plays an important role in stress response, essentially required for Mycobacterium survival. As concluded by various studies that Obg is crucial for Mycobacterium survival under stress, this structural information may help us in designing novel and potential inhibitors against resistant Mycobacterium strains.

Children aged two to four are able to scribble and draw using a smartphone app.

AIM: Drawing apps are an attractive medium, and this study assessed the fluency and skills with which children aged two to four drew using an app on a smartphone. METHODS: We provided 90 children between two and four years of age with a drawing app at a playschool in an upper-class suburb in New Delhi in November 2016. The app allowed them to draw by sliding their finger on the screen and selecting the drawing colour from a palette. The children were given five minutes to draw with the app. RESULTS: All the children were able to draw with the app. The children aged two and three were in the scribbling stage. The children aged two typically drew zigzag lines with a single colour, while the children aged three were able to draw distinct lines and shapes with multiple colours. The children aged four had moved from the scribbling stage to the preschematic stage. They drew multicoloured figures identifiable as real-world objects and were able to explain what they were drawing. CONCLUSION: We found that drawing apps were appropriate for young children aged two to four and may be used by them along with other drawing media.

N-Glycoproteomics of Human Seminal Plasma Glycoproteins.

Seminal plasma aids sperm by inhibiting premature capacitation, helping in the intracervical transport and formation of an oviductal sperm reservoir, all of which appear to be important in the fertilization process. Epitopes such as Lewis x and y are known to be present on seminal plasma glycoproteins, which can modulate the maternal immune response. It is suggested by multiple studies that seminal plasma glycoproteins play, largely undiscovered, important roles in the process of fertilization. We have devised a strategy to analyze glycopeptides from a complex, unknown mixture of protease-digested proteins. This analysis provides identification of the glycoproteins, glycosylation sites, glycan compositions, and proposed structures from the original sample. This strategy has been applied to human seminal plasma total glycoproteins. We have elucidated glycan compositions and proposed structures for 243 glycopeptides belonging to 73 N-glycosylation sites on 50 glycoproteins. The majority of the proposed glycan structures were complex type (83%) followed by high-mannose (10%) and then hybrid (7%). Most of the glycoproteins were either sialylated, fucosylated, or both. Many Lewis x/a and y/b epitopes bearing glycans were found, suggesting immune-modulating epitopes on multiple seminal plasma glycoproteins. The study also shows that large scale N-glycosylation mapping is achievable with current techniques and the depth of the analysis is roughly proportional to the prefractionation and complexity of the sample.

Structural and biochemical characterization of GTP cyclohydrolase II from Helicobacter pylori reveals its redox dependent catalytic activity.

GTP cyclohydrolase II (GCHII), catalyzes the conversion of GTP to 2,5-diamino-6-ß-ribosyl-4(3H)-pyrimidinone-5'-phosphate and has been shown to be essential for pathogens. Here we describe the biochemical, kinetic and structural characterization of GCHII from Helicobacter pylori (hGCHII). The crystal structure of hGCHII, unlike other GCHII structures, revealed that cysteines at the active site existed in oxidized state forming two disulfide bonds and lacked Zn(2+) that was shown to be indispensable for catalytic activity in other species. However, incubation of hGCHII with hydrogen peroxide, an oxidizing agent, followed by PAR-assay showed that Zn(2+) was intrinsically present, indicating that all cysteines at the catalytic site remained in reduced state. Moreover, site directed mutagenesis of catalytic site cysteines revealed that only three, out of four cysteines were essential for hGCHII activity. These results, though, indicated that hGCHII crystallized in oxidized form, the expulsion of Zn(2+) upon oxidation of catalytic cysteines revealed its ability to act in response to the redox environment. Exploring further, incubation of hGCHII with reversible thiol modifying agent S-methyl-methane-thiosulfonate resulted in loss of GCHII activity due to oxidation of its cysteine residues as revealed by mass spectrometry studies. However, addition of reducing agent DTT partially restored the hGCHII catalytic activity. Taken together, these results demonstrate that hGCHII can regulate its catalytic activity depending on the redox environment, a function hitherto unknown for GCHII.

Development of peptide inhibitor as a therapeutic agent against head and neck squamous cell carcinoma (HNSCC) targeting p38alpha MAP kinase.

BACKGROUND: The p38alpha MAP kinase pathway is involved in inflammation, cell differentiation, growth, apoptosis and production of pro-inflammatory cytokines TNF-alpha and IL-1beta. The overproduction of these cytokines plays an important role in cancer. The aim of this work was to design a peptide inhibitor on the basis of structural information of the active site of p38alpha. METHODS: A tetrapeptide, VWCS as p38alpha inhibitor was designed on the basis of structural information of the ATP binding site by molecular modeling. The inhibition study of peptide with p38alpha was performed by ELISA, binding study by Surface Plasmon Resonance and anti-proliferative assays by MTT and flow cytometry. RESULTS: The percentage inhibition of designed VWCS against pure p38alpha protein and serum of HNSCC patients was 70.30 and 71.5%, respectively. The biochemical assay demonstrated the K(D) and IC50 of the selective peptide as 7.22 x 10(-9) M and 20.08 nM, respectively. The VWCS as inhibitor significantly reduced viability of oral cancer KB cell line with an IC50 value of 10 microM and induced apoptosis by activating Caspase 3 and 7. CONCLUSIONS: VWCS efficiently interacted at the ATP binding pocket of p38alpha with high potency and can be used as a potent inhibitor in case of HNSCC. GENERAL SIGNIFICANCE: VWCS can act as an anticancer agent as it potentially inhibits the cell growth and induces apoptosis in oral cancer cell-line in a dose as well as time dependent manner. Hence, p38alpha MAP kinase inhibitor can be a potential therapeutic agent for human oral cancer.

Granulomatous invasive aspergillosis of paranasal sinuses masquerading as actinomycosis and review of published literature.

Cutaneous aspergillosis is a common systemic mycosis affecting immunosuppressed patients. Here, we describe a novel morphological type of cutaneous aspergillosis in a young immunocompetent woman who presented with a chronic history of multiple nodules and discharging sinuses over left side of the face, mimicking cervicofacial actinomycosis. Skin biopsy showed granulomatous inflammation, and of septate fungal hyphae with acute-angled branching, morphologically resembling Aspergillus. This was confirmed on fungal culture as Aspergillus flavus.

Quantitative Proteomics of COVID-19 Recovered Patients Identifies Long-Term Changes in Sperm Proteins Leading to Cellular Stress in Spermatozoa.

Following an initial recovery, COVID-19 survivors struggle with a spectrum of persistent medical complications, including fatigue, breathlessness, weight loss, hair loss, and attention deficits. Additionally, there is growing evidence of adverse effects of COVID-19 on the male reproductive system. This investigation seeks to understand the long-term ramifications on male fertility by examining hormonal profiles, semen parameters, and sperm proteome of recovered COVID-19 patients compared to controls. The serum hormone profiles between the two groups showed minimal variations except for prolactin, cortisol, and testosterone levels. Testosterone levels were slightly lower, while prolactin and cortisol were elevated in COVID-19 cases compared to controls. Though semen parameters exhibited no significant disparities between the COVID-19 and control groups, quantitative proteomics analysis revealed changes in sperm proteins. It identified 190 differentially expressed proteins, of which 161 were upregulated and 29 downregulated in COVID-19 cases. Western blotting analysis validated the differential expression of serpin B4 and calpain 2. Bioinformatics analysis signifies cellular stress in the spermatozoa of COVID-19 recovered patients and thus, SOD and MDA levels in semen were measured. MDA levels were found to be significantly elevated, indicating lipid peroxidation in COVID-19 samples. While the effects of COVID-19 on semen parameters may exhibit a potential for reversal within a short duration, the alterations it inflicts on sperm proteome are persisting consequences on male fertility. This study paves the path for further research and emphasizes the significance of comprehending the complex molecular processes underlying the long-term consequences of COVID-19 on male reproductive health.

The crystal structure reveals the molecular mechanism of bifunctional 3,4-dihydroxy-2-butanone 4-phosphate synthase/GTP cyclohydrolase II (Rv1415) from Mycobacterium tuberculosis.

The enzymes 3,4-dihydroxy-2-butanone 4-phosphate synthase (DHBPS) and GTP cyclohydrolase II (GCHII) catalyze the initial steps of both branches of the bacterial riboflavin-biosynthesis pathway. The structures and molecular mechanisms of DHBPS and GCHII as separate polypeptides are known; however, their organization and molecular mechanism as a bifunctional enzyme are unknown to date. Here, the crystal structure of an essential bifunctional DHBPS/GCHII enzyme from Mycobacterium tuberculosis (Mtb-ribA2) is reported at 3.0 Šresolution. The crystal structure revealed two conformationally different molecules of Mtb-ribA2 in the asymmetric unit that form a dimer via their GCHII domains. Interestingly, analysis of the crystal packing revealed a long `helical-like oligomer' formed by DHBPS and GCHII functional homodimers, thus generating an `open-ended' unit-cell lattice. However, size-exclusion chromatography studies suggest that Mtb-ribA2 exists as a dimer in solution. To understand the discrepancy between the oligomerization observed in solution and in the crystal structure, the DHBPS (Mtb-DHBPS) and GCHII (Mtb-GCHII) domains of Mtb-ribA2 have been cloned, expressed and purified as His-tagged proteins. Size-exclusion chromatography studies indicated that Mtb-GCHII is a dimer while Mtb-DHBPS exists as a monomer in solution. Moreover, kinetic studies revealed that the GCHII activities of Mtb-ribA2 and Mtb-GCHII are similar, while the DHBPS activity of Mtb-ribA2 is much higher than that of Mtb-DHBPS alone. Taken together, the results strongly suggest that Mtb-ribA2 exists as a dimer formed through its GCHII domains and requires full-length Mtb-ribA2 for optimal DHBPS activity.

Isolated corymbose collagenoma responding to intralesional triamcinolone acetonide and hyaluronidase injections.

Collagenomas are connective tissue nevi with circumscribed hamartomatous proliferation of collagen. Due to their benign nature and lack of any simple medical treatment, they are most often left untreated. We present a case of isolated corymbose collagenoma, a distinct morphological variant not described hitherto that was successfully treated with intralesional injections of combination of triamcinolone acetonide and hyaluronidase.