Anticancer and antimicrobial potential of enterocin 12a from Enterococcus faecium.

BACKGROUND: Increase in the number of infections caused by Gram-negative bacteria in neutropenic cancer patients has prompted the search for novel therapeutic agents having dual anticancer and antimicrobial properties. Bacteriocins are cationic proteins of prokaryotic origin that have emerged as one of the most promising alternative antimicrobial agents with applications as food preservatives and therapeutic agents. Apart from their antimicrobial activities, bacteriocins are also being explored for their anticancer potential. RESULTS: In this study, a broad-spectrum, cell membrane-permeabilizing enterocin with a molecular weight of 65 kDa was purified and characterized from the culture supernatant of vaginal Enterococcus faecium 12a. Enterocin 12a inhibited multidrug-resistant strains of various Gram-negative pathogens such as Salmonella enterica, Shigella flexneri, Vibrio cholerae, Escherichia coli and Gram-positive, Listeria monocytogenes, but had no activities against different strains of gut lactobacilli. The mass spectrometric analysis showed that the enterocin 12a shared partial homology with 4Fe-4S domain-containing redox protein of E. faecalis R712. Further, enterocin 12a selectively inhibited the proliferation of various human cancer cell lines in a dose-dependent manner but not that of normal human peripheral blood mononuclear cells. Enterocin 12a-treated cancer cells showed apoptosis-like morphological changes. CONCLUSION: Enterocin 12a is a novel bacteriocin that has anticancer properties against human cell lines and negligible activity towards non-malignant cells. Therefore, it should be further evaluated for its anticancer potential in animal models.

Putative functional non-coding polymorphisms in SELP significantly modulate sP-selectin levels, arterial stiffness and type 2 diabetes mellitus susceptibility.

BACKGROUND: P-selectin, encoded by SELP, has been implicated as an important molecule in the development of arterial stiffness, consequently leading to vascular complications in T2DM. SELP polymorphisms and increased levels of soluble P-selectin (sP-selectin) have been shown to be associated with several inflammatory diseases. The present work was designed to assess nine putative functional non-coding SELP variants in relation to sP-selectin levels and arterial stiffness in T2DM. METHODS: The genetic distribution of rs3917655, rs3917657, rs3917739, rs2235302, rs3917843 was determined by restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR). Genotyping of rs3917779 was performed by tetra primer amplification-refractory mutation system (ARMS)- PCR. Three SNPs i.e. rs3917853, rs3917854, rs3917855 were genotyped by Sanger sequencing. Construction of haplotypes was performed using PHASE software. The data thus obtained was analyzed by appropriate statistical tools. RESULTS: Two non-coding variants i.e. rs3917657 and rs3917854 of SELP were found to be associated with 2 and 1.7 -fold risk of disease development respectively. However, one non-coding variant rs2235302 was found to provide protection against disease development. Furthermore, variant allele of rs3917854 in T2DM patients was found to be associated with 2.07-fold very high vascular risk. Non-coding haplotype GCAGGCCGC was conferring 4.14-fold risk of disease development. Furthermore, overall sP-selectin levels were higher in T2DM patients when segregated according to genotypes as well as haplotypes. Significant genotype- phenotype correlation was observed for rs3917655 as well as rs3917739 variant in patients and for rs3917854 in controls. In vascular risk categories, a significant genotype- phenotype correlation was observed for rs3917655 and rs2235302. Furthermore, patients with CCGGGCCGC haplotype in high risk category were observed with higher levels of sP-selectin as compared to other haplotypes (p < 0.05). CONCLUSIONS: Non-coding SELP variants may significantly modulate sP-selectin levels, vascular risk and T2DM susceptibility.

Association of SELP Polymorphisms with Soluble P-Selectin Levels and Vascular Risk in Patients with Type 2 Diabetes Mellitus: A Case-Control Study.

P-selectin, an adhesion molecule, is encoded by SELP and known as biomarker of endothelial as well as platelet dysfunction. SELP polymorphisms (rs6136, rs6127, and rs6125) and raised levels of soluble P-selectin (sP-selectin) have been associated with several disease conditions. The present study was aimed to determine the association of SELP variants and sP-selectin levels as well as vascular risk in Type 2 diabetes mellitus (T2DM) patients. The frequency of rs6136, rs6127, and rs6125 was assessed by restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR). sP-selectin levels were measured using commercially available kits. Haplotypes were constructed using PHASE software. The data obtained from the above-said analyses was subjected to suitable statistical analyses. sP-selectin levels (ng/ml) were significantly higher in patients as compared to controls (p < 0.001). Out of total, 22% of patients were found to have very high vascular risk, 43.2% with high vascular risk, while 34.4% with moderate vascular risk. For both rs6136 and rs6127, frequency of variant allele was found to be significantly higher in patients as compared to controls and accounted for 2.4- and 1.5-fold risk of disease development, respectively. CAG was found to be associated with 4.5-fold risk towards disease development. In contrast, AGG was conferring the protective effect. Significantly high sP-levels were observed in patients with homozygous wild genotype of rs6136, all genotypes of rs6127, and heterozygous genotype of rs6125 as compared to respective controls. Significant difference was observed in P-selectin levels within moderate-risk category for rs6136. When compared between the categories, significant difference was observed for rs6136 and rs6127. Furthermore, patients with haplotypes AAA, AGA, and AGG were found to have significantly high sP-selectin levels as compared to controls. Significant difference in sP-selectin levels was observed within very high-risk as well as high-risk category. When compared between the categories, significant difference was observed for AGA and AGG haplotypes. The studied polymorphisms of SELP have shown significant association with sP-selectin levels as well as vascular risk in T2DM patients.

Endothelial dysfunction and platelet hyperactivity in type 2 diabetes mellitus: molecular insights and therapeutic strategies.

The incidence and prevalence of diabetes mellitus is rapidly increasing worldwide at an alarming rate. Type 2 diabetes mellitus (T2DM) is the most prevalent form of diabetes, accounting for approximately 90-95% of the total diabetes cases worldwide. Besides affecting the ability of body to use glucose, it is associated with micro-vascular and macro-vascular complications. Augmented atherosclerosis is documented to be the key factor leading to vascular complications in T2DM patients. The metabolic milieu of T2DM, including insulin resistance, hyperglycemia and release of excess free fatty acids, along with other metabolic abnormalities affects vascular wall by a series of events including endothelial dysfunction, platelet hyperactivity, oxidative stress and low-grade inflammation. Activation of these events further enhances vasoconstriction and promotes thrombus formation, ultimately resulting in the development of atherosclerosis. All these evidences are supported by the clinical trials reporting the importance of endothelial dysfunction and platelet hyperactivity in the pathogenesis of atherosclerotic vascular complications. In this review, an attempt has been made to comprehensively compile updated information available in context of endothelial and platelet dysfunction in T2DM.

Targeting dendritic cell-specific TNFR2 improves skin and joint inflammation by inhibiting IL-12/ IFN-γ pathways in a mouse model of psoriatic arthritis.

Psoriasis (PsO) and Psoriatic arthritis (PsA) are immune-mediated inflammatory diseases affecting the skin and joints. Approximately, 30% of patients with PsO develop PsA over time with both conditions being associated with elevated tumor necrosis factor-alpha (TNF-α) expression. TNF-α mediates its effect through two membrane receptors, TNFR1 and TNFR2. While current TNF-α-neutralizing agents, targeting both TNFR1 and TNFR2 receptors, constitute the primary treatment for psoriatic diseases, their long-term use is limited due to an increase in opportunistic infections, tuberculosis reactivation and malignancies likely attributed to TNFR1 inactivation. Recent findings suggest a pivotal role of TNFR2 in psoriatic disease, as evidenced by its amelioration in global TNFR2-knockout (TNFR2KO) mice, but not in TNFR1KO mice. The diminished disease phenotype in TNFR2KO mice is accompanied by a decrease in DC populations. However, the specific contribution of TNFR2 in dendritic cells (DCs) remains unclear. Here, utilizing a mannan-oligosaccharide (MOS)-induced PsA model, we demonstrate a significant reduction in PsA-like skin scaling and joint inflammation in dendritic cell-specific TNFR2 knockout mice (DC-TNFR2KO). Notably, MOS treatment in control mice (TNFR2 fl/fl) led to an increase in conventional type 1 dendritic cells (cDC1) population in the spleen, a response inhibited in DC-TNFR2KO mice. Furthermore, DC-TNFR2KO mice exhibited reduced levels of interleukin-12 (IL-12), a Th1 cell activator, as well as diminished Th1 cells, and interferon-gamma (IFN-γ) levels in the serum compared to controls following MOS stimulation. In summary, our study provides compelling evidence supporting the role of TNFR2 in promoting PsA-like inflammation through cDC1/Th1 activation pathways.

GRAPPA 2022 Trainee Symposium: A Summary of Oral and Poster Presentations.

One of the highlights of the 2022 Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) annual meeting was the trainee symposium. Dermatology and rheumatology trainees presented their research related to psoriasis and psoriatic arthritis. This report briefly reviews 5 oral presentations and 15 posters that were selected for this annual meeting. Topics include basic/translational, clinical, and outcomes research reflecting the spectrum of GRAPPA's effort and influence nationally and internationally in the area of psoriatic diseases.

Therapeutic Modalities in Head-Neck Neoplasia and Prognostication: A Review.

Neoplasia of the head and the neck necessitates intervention, surgical or otherwise, as the site and stage of the pathology may dictate. The various therapeutic modalities employed and prognosis has been reviewed.

2021 GRAPPA Trainee Symposium: A Summary of Oral and Poster Presentations.

The Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) held a trainee symposium at its 2021 virtual meeting. Dermatology and rheumatology trainees presented their work on psoriasis and psoriatic arthritis (PsA). This report briefly reviews 5 oral presentations: prediction of cardiovascular events in psoriatic disease (PsD), correlation between spine abnormalities and clinical findings, biomechanical stress as a trigger for PsA, differences in DNA methylation among twins with PsD, and critical proteins associated with induction of PsD. In addition, we highlight 22 posters broadly discussing clinical and molecular implications of PsD.

TNFR2 Depletion Reduces Psoriatic Inflammation in Mice by Downregulating Specific Dendritic Cell Populations in Lymph Nodes and Inhibiting IL-23/IL-17 Pathways.

TNF-α, a proinflammatory cytokine, is a crucial mediator of psoriasis pathogenesis. TNF-α functions by activating TNFR1 and TNFR2. Anti-TNF drugs that neutralize TNF-α, thus blocking the activation of TNFR1 and TNFR2, have been proven highly therapeutic in psoriatic diseases. TNF-α also plays an important role in host defense; thus, anti-TNF therapy can cause potentially serious adverse effects, including opportunistic infections and latent tuberculosis reactivation. These adverse effects are attributed to TNFR1 inactivation. Therefore, understanding the relative contributions of TNFR1 and TNFR2 has clinical implications in mitigating psoriasis versus global TNF-α blockade. We found a significant reduction in psoriasis lesions as measured by epidermal hyperplasia, characteristic gross skin lesion, and IL-23 or IL-17A levels in Tnfr2-knockout but not in Tnfr1-knockout mice in the imiquimod psoriasis model. Furthermore, imiquimod-mediated increase in the myeloid dendritic cells, TNF/inducible nitric oxide synthase‒producing dendritic cells, and IL-23 expression in the draining lymph nodes were dependent on TNFR2 but not on TNFR1. Together, our results support that psoriatic inflammation is not dependent on TNFR1 activity but is driven by a TNFR2-dependent IL-23/IL-17 pathway activation. Thus, targeting the TNFR2 pathway may emerge as a potential next-generation therapeutic approach for psoriatic diseases.

Preliminary studies of the effect of doping of chromium oxide in SiO2-CaO-P2O5 bioceramics for bone regeneration applications.

Bioceramics of composition xCr2O3∙(43-x) CaO∙42SiO2∙15P2O5 (x varying from 0 to 8 mol%) have been synthesized in the laboratory by using sol-gel technique. The morphology and structure has been determined by using Powder X-ray Diffraction, Fourier Transform Infrared and Raman spectroscopy and Field Emission Scanning Electron Microscopy. The in vitro bio mineralization behavior has been assessed by immersion in simulated body fluid for 7 days. The results obtained in our studies have indicated excellent hydroxyapatite formation ability of our samples. Drug delivery property of synthesized samples has been checked by using UV-spectroscopy of antibiotic 'gentamicin'. The in vitro drug release profile was fitted best in the Higuchi model with the highest value of coefficient of determination (R2 = 0.9970). Antimicrobial properties have been evaluated from minimum inhibitory concentration and time kill assay values. The cellular response has been investigated by using human osteosarcoma MG 63 cell line. Also to check charge on the synthesized samples, Zeta potential studies have been conducted and it has been observed that samples carry negative charge when immersed in simulated body fluid. Negative surface charge provide suitable environment for cell adhesion and proliferation. Experiments have been undertaken to explore suitable composition with an objective of development of suitable implant material for bone regeneration applications.

Silver derivatives of multi-donor heterocyclic thioamides as antimicrobial/anticancer agents: unusual bio-activity against methicillin resistant S. aureus, S. epidermidis, and E. faecalis and human bone cancer MG63 cell line.

The basic aim of this study pertains to the synthesis of silver nitrate complexes and the study of their antimicrobial and anticancer bio-activity. A series of new silver(i) derivatives of N-substituted-imidazolidine-2-thiones (L-NR, R = Et, Pr n , Bu n , Ph), purine-6-thione (purSH2), 2-thiouracil (tucH2), pyrimidine-2-thione (pymSH) and pyridine-2-thione (pySH) of composition [Ag(S-L-NR)(PPh3)2(ONO2)] {R = Et (1), Pr n (2), Bu n (3), Ph (4)}, [Ag2(N,S-purSH2)2(µ-dppm)2](NO3)2·2H2O (5) (dppm = Ph2P-CH2-PPh2), [Ag(L)(PPh3)2](NO3) {L = N,S-purSH2 (6); S-tucH2 (7)}, [Ag(N,S-pymS)(PPh3)2](CH3OH) (8), and [Ag(N,S-pyS)(PPh3)2] (9) have been synthesized and structurally characterized. These new and some previously reported complexes {[Ag2(L-NH)4(PPh3)2](NO3)2 (10), [Ag(L-NMe)2(PPh3)](NO3) (11), and [Ag(S-bzimSH)2(PPh3)2](OAc) (12), L-NH = 1,3-imidazolidine-2-thione; L-NMe = 1-methyl-3-imidazolidine-2-thione and bzimSH2 = benzimidazoline-2-thione)} have shown moderate to high anti-microbial activity against Gram positive bacteria, namely methicillin resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus (MTCC 740), and Gram negative bacteria, namely Staphylococcus epidermidis (MTCC 435), Enterococcus faecalis (MTCC 439), Shigella flexneri (MTCC 1457) and a yeast Candida albicans (MTCC 22). These complexes have also been found to be bio-safe as studied using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay. The anti-tumor study of silver complexes against human osteosarcoma cell line (MG63) has shown IC50 values in the range of 6-33 µM.

Proteinaceous Secretory Metabolites of Probiotic Human Commensal Enterococcus hirae 20c, E. faecium 12a and L12b as Antiproliferative Agents Against Cancer Cell Lines.

Cancer is the second leading cause of death worldwide and its incidence is expected to grow by almost 70% in the coming 2 decades. Recent microbiome studies in cancer mice models have shown that certain commensal bacteria play protective roles against cancer. Thus, the use of commensal microflora having anticancer activities for the treatment of cancer appears to be an attractive alternative therapeutic strategy. Lactic acid bacteria (LAB) form an integral component of commensal microflora in healthy individuals. As the vaginal ecosystem is enriched in LAB genera, we screened the vaginal LAB microflora of healthy women for their anti-proliferative abilities against various human cancer cell lines. The secreted metabolites of three enterococcal strains, Enterococcus hirae 20c, Enterococcus faecium 12a and L12b, out of 92 LAB isolates selectively inhibited the in vitro proliferation of various human cancer cell lines in a dose-dependent manner but had no activity against normal human peripheral blood monocytes. Further, proteinase K-treatment of the cell-free supernatant (CS) of all the three enterococci abrogated their anti-proliferative abilities, thereby showing the proteinaceous nature of the secreted metabolites in the CS. The microscopic examination of the cell lines showed that CS-treatment induced apoptosis-like morphological changes in the cancer cells. Further, the probiotic characters of the strains were studied, which showed that all the three strains had broad spectrum antimicrobial activities against various Gram-positive and Gram-negative pathogens, including Mycobacterium smegmatis. All the strains tolerated the gastric acidity and bile juice treatments, and had strong adhesive abilities to the colonic epithelial cell line HCT-15. Furthermore, none of the strains had any known secreted virulence factors or harbored virulence genes. This preliminary study highlights an important functional role of the commensal probiotic enterococcal strains E. hirae and E. faecium for the first time by demonstrating their anticancer properties that should be further tested in the in vivo mammalian models.

Scaffolds of hydroxyl apatite nanoparticles disseminated in 1, 6-diisocyanatohexane-extended poly(1, 4-butylene succinate)/poly(methyl methacrylate) for bone tissue engineering.

Poly(1, 4-butyl succinate) extended 1, 6-diisocyanatohexane (PBSu-DCH) polymers and Polymethylmethacrylate (PMMA) scaffolds decorated with nano hydroxyl apatite have been prepared and characterized for regeneration of bone in cranio-maxillofacial region. Synthesized scaffolds revealed good response in bone regeneration and excellent cell viability in comparison to commercial available glass plate, which lead to better proliferation of MG-63 cell lines. Additionally, they demonstrate high porosity and excellent water retention ability. Moreover, controlled degradation (in pH=7.4) and sustained drug release in pH (4.5 and 7.4) are advantages of these scaffolds to serve as delivery vehicles for therapeutic drugs. Samples also provide the protection against Escherichia coli and Methicillin Resistant Staphylococcus aureus microorganisms which can be helpful for quick recovery of the patient. In-vitro inflammatory response has been assessed via adsorption of human plasma/serum proteins on the surface of the scaffolds. Results suggest that prepared scaffolds have good bone regeneration ability and provide friendly environment for the cell growth with the additional advantage of protection of the surrounding tissues from microbial infection. With all these features, it is speculated that these scaffolds will have wide utility in the area of tissue engineering and regenerative medicine.