Biology subjects

Senthilkumar, R.

Publications and source records attributed to Senthilkumar, R..

7 recordsLinked to original sources

bioRxiv2023

A hybrid combination of in vitro cultured buccal mucosal cells using two different methodologies, complementing each other in successfully repairing a stricture-inflicted human male urethral epithelium

BackgroundAutologous buccal mucosal tissue derived cell transplantation techniques in repairing a stricture inflicted male urethral epithelium have been evolving. There was not much of clarity on the cell type, in vitro culture methods and the mode of transplantation, until we reported our buccal epithelium expanded and encapsulated in scaffold-hybrid approach to urethral stricture (BEES-HAUS) clinical study yielding a successful engraftment and repair with a long-term patency. We herein report with technical clarity on the advantages of mixing two-dimensional (2D) monolayer cultured fibroblast like cells and three dimensional (3D) thermo-reversible gelation polymer (TGP) cultured cells; the former secreting IGF-1, a cytokine known for its healing effects and the latter expressing epithelial surface markers in flow cytometry, both sourced from human buccal tissue, together transplanted using TGP as a carrier. MethodsHuman buccal tissues (n=22) redundant after urethroplasty surgery was used after informed consent and IEC approval. They were enzyme digested, divided into two portions; one was cultured as monolayer method (2D) and the other in 3D in TGP. Flowcytometry and quantification of IGF-1 in cell culture supernatant through the culture period were undertaken. ResultsIn flowcytometry, the cells on day 0, lacked AE1/AE3 - pancytokeratin expression indicative of epithelial phenotype of culture, which progressively increased in the 3D-TGP group, during invitro culture of up to 21 days. The 2D showed expression of only fibroblasts like cells that were negative for AE1/AE3 but positive for CD140b. IGF-1 secretion was significantly higher in 2D cultures than in 3D-TGP (p-value < 0.05). ConclusionThe 3D- TGP cultured cells of epithelial nature and the 2D cultured fibroblast like cells secreting IGF-1, together when transplanted using TGP scaffold as a carrier, adapted to a hostile in vivo milieu after releasing the fibrous strands with urethrotomy, successfully engrafted and repaired a stricture-inflicted male urethral epithelium in the BEES-HAUS procedure. While this hybrid combination of cells are considered to have potential in managing other epithelial damages, further research of such hybrid combination and their behaviour in disease affected environments may help to expand this solution in regenerating and repairing other tissues and organs as well. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=123 SRC="FIGDIR/small/555240v1_ufig1.gif" ALT="Figure 1"> View larger version (70K): org.highwire.dtl.DTLVardef@1e00b7eorg.highwire.dtl.DTLVardef@12886aorg.highwire.dtl.DTLVardef@b9fbb3org.highwire.dtl.DTLVardef@3bd247_HPS_FORMAT_FIGEXP M_FIG C_FIG Illustration of pathogenesis of urethral stricture and the contribution of the hybrid combination of two-dimensional (2D) and three dimensional, 3D-TGP (Thermo-reversible gelation polymer) cultured cells to the successful repair of the stricture-inflicted male urethral epithelium in the buccal epithelium expanded and encapsulated in scaffold-hybrid approach to urethral stricture (BEES-HAUS) technique.

synthetic biology↗

bioRxiv2023

Enhanced muscle regeneration in mdx mice, Duchenne muscular dystrophy animal model, proven by CD44 & MYH3 expression, on oral feeding of N-163 strain of Aureobasidium Pullulans produced B-Glucan

IntroductionDuchenne muscular dystrophy (DMD) is a rare genetic disease, causing muscle degeneration due to lack of dystrophin with inadequate muscle regeneration culminating in muscle dysfunction. The N-163 strain of Aureobasidium Pullulans produced Beta-1,3-1,6-glucan (Neu REFIX) reported to be safe with anti-inflammatory and anti-fibrotic efficacy earlier, herein we evaluated its effects on muscle regeneration in mdx mice. MethodsForty-five mice in three groups (n=15 each): Group 1 (normal), Group 2 (mdx control), and Group 3 (mdx fed Neu REFIX) were evaluated for 45 days. IGF-1, Dystrophin, CD44 and MYH3 in diaphragm, plasma and skeletal muscle were evaluated by ELISA and immunohistochemistry. ResultsMean IGF-1 expression was 20.32% and 16.27% higher in plasma (p = 0.03) and diaphragm respectively in Neu-REFIX group. Mean dystrophin was higher in Neu-REFIX group by 70.3% and 4.7% in diaphragm and plasma respectively than control. H-score intensity of CD44+ was >2.0 with an MYH3-positivity 20% higher in Neu-REFIX than control. ConclusionOral administration of Neu REFIX was safe. Significantly enhanced plasma IGF-1 beside increased Dystrophin, MYH3 and CD44, proving a restoration of muscle regeneration and differentiation, especially in diaphragm, makes us recommend it as a disease modifying adjuvant in both early and advanced stages of DMD.

genetics↗

bioRxiv2022

Resolution of fibrosis in mdx dystrophic mouse after oral consumption of N-163 strain of Aureobasidium pullulans produced biological response modifier β-glucan (BRMG)`

Recent advances in the management of Duchenne Muscular Dystrophy (DMD), such as exon skipping therapy, have reached a clinical stage, and although gene therapy is in clinical trials, the outcome at its best is still considered suboptimal to yield clinically discernible progress. In this study, we evaluated a novel N-163 strain of Aureobasidium pullulans produced biological response modifier {beta}-glucan (BRMG) for its potential as an adjuvant to slow down the progression of the disease by anti-inflammatory and anti-fibrotic effects. This N-163 {beta}-glucan is a safe and orally consumable food supplement with similar effects that have been previously proven in pre-clinical studies of organ fibrosis, and their beneficial effects have been proven in DMD clinical studies through blood parameters as well. In this study, 45 mice in the three groups, 15 each in a group; Gr. 1 normal mice, Gr.2 mdx mice as vehicle, and Gr.3 mdx mice administered the N-163 strain produced {beta}-glucan for 45 days. Blood biochemical parameters, body weight, muscle weight, inflammation score, and fibrosis score were evaluated using H&E and Massons trichrome staining. The N-163 {beta}-glucan group showed a significant decrease in the plasma ALT, AST, and LDH levels (126 {+/-} 69, 634 {+/-} 371, 3335 {+/-} 1258 U/l) compared with the vehicle group (177 {+/-} 27 U/l, 912 {+/-} 126 U/l, 4186 {+/-} 398 U/l). Plasma TGF-{beta} levels increased, and plasma IL-13 levels decreased in the N-163 group. The inflammation score of HE-stained muscle sections in the N-163 group (1.5 {+/-} 0.8) was lower than that in the vehicle group (2.0 {+/-} 0.8). The percentage of centrally nucleated fibres (CNF) evaluated by Massons trichrome staining was 0 in the normal group, while it increased to 80% in the vehicle group and 76.8% in the N-163 group. The N-163 {beta}-glucan group (24.22 {+/-} 4.80) showed a significant decrease in the fibrosis area (Massons trichrome-positive area). The N-163 {beta}-glucan thus, demonstrated its anti-fibrotic effect in this study. Considering their safety and easy oral consumption, this BRMG could be worth large multicentre clinical studies as adjuvant in slowing down the progress of DMD.

genetics↗

bioRxiv2022

Two unique biological response-modifier glucans beneficially regulating gut microbiota and faecal metabolome in a non-alcoholic steatohepatitis animal model, with potential for applications in human health and disease

ObjectiveThe gut microbiome and its metabolites, influenced by age and stress, reflect the metabolism and immune systems health. We assessed the gut microbiota and faecal metabolome in a Stelic Animal Model of non-alcoholic steatohepatitis (NASH). DesignThis model was subjected to the following treatments: reverse osmosis water, AFO-202, N-163, AFO-202+N-163, and telmisartan. Faecal samples were collected at 6 weeks and 9 weeks of age. The gut microbiome was analysed using 16S ribosomal RNA sequence acquired by next-generation sequencing and the faecal metabolome using gas chromatography-mass spectrometry. ResultsThe gut microbial diversity increased greatly in the AFO-202+N-163 group. Post-intervention, the abundance of Firmicutes decreased, while that of Bacteroides increased and was the highest in the AFO-202+N-163 group. The decrease in the Enterobacteria and other Firmicutes abundance and in the Turicibacter and Bilophila abundance was the highest in the AFO-202 and N-163 groups, respectively. The Lactobacillus abundance increased the most in the AFO-202+N-163 group. The faecal metabolites spermidine and tryptophan, beneficial against inflammation and NASH, respectively, were greatly increased in the N-163 group. Succinic acid, beneficial in neurodevelopmental and neurodegenerative diseases, increased in the AFO-202 group. Decrease in fructose was the highest in the AFO-202 group. Leucine and phenylalanine decreased, whereas ornithine, which is beneficial against chronic immune-metabolic-inflammatory pathologies, increased in the AFO-202+N-163 group. ConclusionAFO-202 treatment in mice is beneficial against neurodevelopmental and neurodegenerative diseases and has prophylactic potential against metabolic conditions. N-163 treatment has anti-inflammatory effects against organ fibrosis and neuroinflammatory conditions. In combination, they present anticancer activity. Key messagesO_LIThe influence of gut microbiome on fecal metabolome and their association to several diseases is already known. C_LIO_LIThis study proves the efficacy of 1,3-1,6 beta glucans with pre-biotic potentials, beneficially influencing both gut microbiome and metabolome. C_LIO_LIThese results recommends for an in-depth exploration of relationship among pre-biotics, gut microbiome and gut-multi-organ axes on the fundamentals of disease onset. C_LIO_LIHidden prophylactic and therapeutic solutions to non-contagious diseases with Aureobasidium pullulans produced 1,3-1,6 beta glucans may be unveiled. C_LI

microbiology↗

bioRxiv2021

Beneficial effects of the AFO-202 and N-163 strains of Aureobasidium pullulans produced 1,3-1,6 beta glucans on non-esterified fatty acid levels in obese diabetic KKAy mice: A comparative study

Obesity, metabolic syndrome, associated lipotoxicity and its cascade of events contribute to the majority of the burden related to non-communicable diseases globally. Preventive lifestyle changes aside, several beneficial effects have been reported in type II diabetes mellitus and dyslipidaemia patients with biological response modifier glucans (BRMG) produced as an exopolysaccharide by Aureobasidium pullulans. In this study, we compared two strains (AFO-202 and N-163) that produce beta glucans in alleviating lipotoxicity. This study was performed in obese diabetic mice model of KK-Ay mice, in four groups with six subjects in each group - Group 1: sacrificed on Day 0 for baseline values; Group 2: control (drinking water); Group 3: AFO-202 beta glucan--200 mg/kg/day; Group 4: N-163 beta glucan--300 mg/kg/day. The animals in groups 2-4 had the test solutions administered by gavage once daily for 28 consecutive days. Biochemical analyses were conducted of blood glucose, triglycerides, total cholesterol, LDL cholesterol, HDL cholesterol and non-esterified fatty acids (NEFA). Group 4 (N-163) had the lowest NEFA levels, as compared to the other groups, and marginally decreased triglyceride levels. The groups had no significant differences in blood glucose, HbA1c, triglycerides, or LDL and HDL cholesterol. N-163 produced by A. pullulans decreased NEFA in a diabetic mice model in 28 days. These results, although modest, warrant further in-depth research into lipotoxicity and associated inflammatory cascades in both healthy and disease affected subjects to develop novel strategies for prevention and management.

pharmacology and toxicology↗

bioRxiv2021

An efficient polymer cocktail-based transportation method for cartilage tissue, yielding chondrocytes with enhanced hyaline cartilage expression during in vitro culturing

Chondrocytes are used in cell-based therapies such as autologous chondrocyte implantation (ACI) and matrix-associated cartilage implantation (MACI). To transport the cartilage tissue to the laboratory for in vitro culturing, phosphate-buffered saline (PBS), Euro-Collins solution (ECS) and Dulbeccos Modified Eagles Medium (DMEM) are commonly employed at 4-8 {degrees}C. In this study, eight samples of human cartilage biopsy tissues from elderly patients with severe osteoarthritis undergoing arthroscopy, which would otherwise have been discarded, were used. The cartilage tissue samples were compared to assess the cell yield between two transportation groups: i) a thermo-reversible gelation polymer (TGP) based method without cool preservation (~25 {degrees}C) and ii) ECS transport at 4 {degrees}C. These samples were subjected to in vitro culture in a two-dimensional (2D) monolayer for two weeks and subsequently in a three-dimensional (3D) TGP scaffold for six weeks. The cell count obtained from the tissues transported in TGP was higher (0.2 million cells) than those transported in ECS (0.08 million cells) both after initial processing and after in vitro culturing for 2 weeks in 2D (18 million cells compared with 10 million cells). In addition, mRNA quantification demonstrated significantly higher expression of Col2a1 and SOX-9 in 3D-TGP cultured cells and lower expression of COL1a1 in RT-PCR, characteristic of the hyaline cartilage phenotype, than in 2D culture. This study confirms that the TGP cocktail is suitable for both the transport of human cartilage tissue and for in vitro culturing to yield better-quality cells for use in regenerative therapies.

cell biology↗

bioRxiv2021

Hepatoprotective effects of Aureobasidium pullulans derived Beta 1,3-1,6 biological response modifier glucans in a STAM- animal model of non-alcoholic steatohepatitis

BackgroundNon-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are highly prevalent conditions characterized by inflammation and fibrosis of the liver which can progress to cirrhosis and hepatocellular carcinoma if left untreated. Lifestyle disorders such as obesity, diabetes and dyslipidaemia predispose to and are associated with the disease progression. Conventional modalities are mainly symptomatic, with no definite solution. Beta glucan-based biological response modifiers are a potential strategy in lieu of their beneficial metabolic effects. Aureobasidium pullulans strains AFO-202 and N-163 beta glucans were evaluated for anti-fibrotic and anti-inflammatory hepatoprotective potentials in a NASH animal model in this study. MethodsIn the STAM murine model of NASH, five groups were studied for eight weeks-- (1) vehicle (RO water), (2) AFO-202 beta glucan; (3) N-163 beta glucan, (4) AFO-202+N-163 beta glucan, and (5) telmisartan (standard pharmacological intervention). Evaluation of biochemical parameters in plasma and hepatic histology including Sirius red staining and F4/80 immunostaining were performed. ResultsAFO-202 beta glucan significantly decreased inflammation-associated hepatic cell ballooning and steatosis. N-163 beta glucan decreased fibrosis and inflammation significantly (p value<0.05). The combination of AFO-202 with N-163 significantly decreased the NAFLD Activity Score (NAS) compared with other groups. ConclusionThis preclinical study supports the potential of N-163 and AFO-202 beta glucans alone or in combination as potential preventive and therapeutic agent(s), for NASH. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

pharmacology and toxicology↗