Evaluation of cell adhesion molecules (LFA-1 and L-selectin) in ankylosing spondylitis patients after treatment with ß-D-mannuronic acid (M2000).

Background & objectives: To examine ß-D-mannuronic acid (M2000) effects on L-selectin shedding and leucocyte function-associated antigen-1 (LFA-1) expression as mechanisms of action of this drug in patients with ankylosing spondylitis (AS). Methods: To investigate the molecular consequences of ß-D-mannuronic acid on L-selectin shedding, flow cytometry method was used. Furthermore, the effect of it on LFA-1 gene expression was analyzed by using quantitative real time (qRT)-PCR technique. Results: The LFA-1 expression in patients with AS was higher than controls (P=0.046). The LFA-1 expression after 12 wk therapy with ß-D-mannuronic acid was meaningfully decreased (P=0.01). After 12 wk treatment with ß-D-mannuronic acid, the frequency of CD62L-expressing CD4+ T cells in patients with AS, was not considerably altered, compared to the patients before therapy (P=0.5). Furthermore, after 12 wk therapy with ß-D-mannuronic acid, L-selectin expression levels on CD4+ T-cells in patients with AS, were not remarkably changed, compared to the expression levels of these in patients before treatment (P=0.2). Interpretation & conclusions: The results of this study for the first time showed that ß-D-mannuronic acid can affect events of adhesion cascade in patients with AS. Moreover, ß-D-mannuronic acid presented as an acceptable benefit to AS patients and could aid in the process of disease management.

Evaluation of the success rate of pit and fissure sealants on first molars: 12 months follow-up study.

INTRODUCTION: Dental caries by far is the most prevalent concern of the preadolescents and adolescents in dental clinics. Despite the provision of local fluoride, the occlusal surfaces of teeth are susceptible to dental caries. Pit and fissure sealant therapy is a preventive method to decrease dental caries in permanent teeth. The present study aimed to evaluate the success of fissure sealant treatments of first molar teeth, at 3, 6 and 12 months follow-ups. MATERIALS AND METHODS: Sixty-five children were randomly selected. The subjects had already received fissure sealants in the department of public health dentistry. Demographic data, including age and gender, sealant failure and the type of failure were recorded in the relevant checklists. Feigal criteria were used to evaluate the success or failure of fissure sealant treatments. RESULTS: Overall success rate was 74.3% for 1 year. Evaluation of the failure rate showed that at the 3-month interval, 20.6% of the sealants exhibited failure (57.1% due to margin discoloration and 42.9% due to lack of margin adaptation). 28.6% of the sealants failed at the 6-month (75% due to marginal discoloration and 25% due to anatomical form) and 41.2% failed at the 12-month interval (57.1% due to marginal discoloration and 42.9% due to the lack of margin adaptation). CONCLUSION: The total failure rate of fissure sealant failures after 1 year was 27.7%. The most frequent reason for the failure of fissure sealants was marginal discoloration.

The osteogenic differentiation of human dental pulp stem cells in alginate-gelatin/Nano-hydroxyapatite microcapsules.

BACKGROUND: Microcapsule is considered as a promising 3D microenvironment for Bone Tissue Engineering (BTE) applications. Microencapsulation of cells in an appropriate scaffold not only protected the cells against excess stress but also promoted cell proliferation and differentiation. Through the current study, we aimed to microcapsulate the human Dental Pulp Stem Cells (hDPSCs) and evaluated the proliferation and osteogenic differentiation of those cells by using MTT assay, qRT-PCR, Alkaline phosphatase, and Alizarine Red S. RESULTS: The SEM results revealed that Alg/Gel microcapsules containing nHA showed a rough and more compact surface morphology in comparison with the Alg/Gel microcapsules. Moreover, the microencapsulation by Alg/Gel/nHA could improve cell proliferation and induce osteogenic differentiation. The cells cultured in the Alg/Gel and Alg/Gel/nHA microcapsules showed 1.4-fold and 1.7-fold activity of BMP-2 gene expression more in comparison with the control group after 21 days. The mentioned amounts for the BMP-2 gene were 2.5-fold and 4-fold more expression for the Alg/Gel and Alg/Gel/nHA microcapsules after 28 days. The nHA, addition to hDPSCs-laden Alg/Gel microcapsule, could up-regulate the bone-related gene expressions of osteocalcin, osteonectin, and RUNX-2 during the 21 and 28 days through the culturing period, too. Calcium deposition and ALP activities of the cells were observed in accordance with the proliferation results as well as the gene expression analysis. CONCLUSION: The present study demonstrated that microencapsulation of the hDPSCs inside the Alg/Gel/nHA hydrogel could be a potential approach for regenerative dentistry in the near future.

Synthesis, characterization, and evaluation of curcumin-loaded endodontic reparative material.

Curcumin (CUR) is an ancient therapeutic agent with remarkable antimicrobial and anti-inflammatory properties. The purpose of the current study was to synthesize and evaluate a curcumin-based reparative endodontic material to reduce infection and inflammation besides the induction of mineralization during the healing of the dentin-pulp complex. Poly-ɛ-caprolactone (PCL)/gelatin (Gel)/CUR scaffold was synthesized and assessed by scanning electron microscopy, Fourier transform infrared spectroscopy, and thermo-gravimetric analysis (TGA). Agar diffusion test was performed against E. coli, A. baumannii, P. aeruginosa, S. aureus, E. faecalis, and S. mutans. Moreover, proliferative, antioxidative, anti-inflammatory, and calcification properties of these scaffolds on human dental pulp stem cells (hDPSCs) were evaluated. The results showed that PCL/Gel/CUR scaffold had antibacterial effects. Also, these CUR-based scaffolds had significant inhibitory effects on the expression of tumor necrosis factor α and DCF from inflamed hDPSCs (p < 0.05). Moreover, the induction of mineralization in hDPSCs significantly increased after seeding on CUR-based scaffolds (p < 0.05). Based on these findings, the investigated CUR-loaded material was fabricated successfully and provided an appropriate structure for the attachment and proliferation of hDPSCs. It was found that these scaffolds had antimicrobial, antioxidant, and anti-inflammatory characteristics and could induce mineralization in hDPSCs, which is essential for healing and repairing the injured dentin-pulp complex.

Effects of guluronic acid, as a new NSAID with immunomodulatory properties on IL-17, RORγt, IL-4 and GATA-3 gene expression in rheumatoid arthritis patients.

Aim: Rheumatoid arthritis (RA) is a prevalent inflammatory, autoimmune diseases characterized by inflammation and destruction of joints. Disease-modifying anti-rheumatic drugs (DMARDs) and biological drugs can have modulatory interference in disease process. In this study, the effect of Guluronic Acid (G2013) as a novel non-steroidal anti-inflammatory drug (NSAID) with immunomodulatory effects was evaluated on IL-17, RORγt, IL-4 and GATA-3 gene expression in RA patients.Methods: Fourteen patients with RA who had an inadequate response to conventional treatments were included in this clinical trial. During this trial, patients were permitted to continue the conventional therapy excluding NSAIDs. G2013 was administered orally at dose of 500 mg twice daily for 12 weeks. The peripheral blood mononuclear cells (PBMCs) were collected before and after treatment to evaluate the gene expression levels of IL-4, GATA-3, IL-17 and RORγt.Results: Primary and secondary efficacy endpoints and Disease Activity Score (DAS) 28 showed an improvement after 12 weeks of treatment. G2013 has a potent efficacy on gene expression of these molecules, so that it could decrease IL-17 and RORγt levels and increase IL-4 and GATA-3 levels after 12 weeks of treatment. Reduction of IL-17 was statistically non-significant whereas for its transcription factor (RORγt) was statistically significant. Moreover, the gene expression results were in accordance with the clinical and preclinical assessments.Conclusion: G2013 as a natural novel drug showed a significant increase on IL-4 and GATA-3 and a significant decrease on RORγt gene expression after 12 weeks oral administration of this drug in RA patients. (Clinical trial identifier: IRCT2016092813739N5).

The effect of electromagnetic fields on survival and proliferation rate of dental pulp stem cells.

Aims: Extremely low-frequency electromagnetic fields (ELF-EMF) can affect biological systems and alter some cell functions like proliferation rate. Dental pulp tissue is known as a source of multipotent stromal stem cells (MSCs), which can be obtained by a less invasive and more available process compared to bone marrow-derived stem cells (BMSCs). This study aimed to consider the effect of ELF-EMF on proliferation rates of human dental pulp stem cells (hDPSCs).Material and methods: ELF-EMF was generated by a system including autotransformer, multi-meter, solenoid coils, teslameter and its probe. The effect of ELF-EMF with the intensity of 0.5 and 1 mT and 50 Hz on the proliferation rate of hDPSCs was assessed in 20 and 40 min per day for 7 days. MTT assay and DAPI test were used to determine the growth and proliferation of DPSCs.Results: Based on MTT, ELF-EMF has maximum effect with the intensity of 1 mT for 20 min/day on the proliferation of hDPSCs. The survival and proliferation rate in all exposure groups were significantly higher than the control group. Based on the data obtained from MTT and DAPI assay, the number of viable cells in the group exposed to 1 mT for 20 min/day was higher than other groups (p < .05).Conclusions: Regarding to the results of this study, 0.5 and 1 mT ELF-EMF can enhance survival and proliferation rates of hDPSCs.

An in vitro assessment for evaluating the efficiency of ß-d-mannuronic acid (M2000) in myelodysplastic syndrome.

ß-d-Mannuronic acid (M2000), a novel non-steroidal anti-inflammatory drug (NSAID) with immunosuppressive properties, has been previously shown to exhibit potential therapeutic effects on autoimmune diseases. Immunosuppression therapy has been a standard approach for myelodysplastic syndrome (MDS) for many years. We evaluated the effect of M2000 on isolated peripheral blood mononuclear cells (PBMCs) from patients with MDS. The PBMCs were isolated from 13 patients with MDS and 13 normal donors. The cells were then treated with low, moderate, and high doses of M2000 and diclofenac as a control group. The level of interleukin (IL)-6, tumor necrosis factor alpha (TNF-α), IL-3, granulocyte colony-stimulating factor (G-CSF) gene expression and the serum level of IL-6 and TNF-α production were evaluated by real-time polymerase chain reaction and enzyme-linked immunosorbent assay methods, respectively. Our findings indicated a significant reduction in the production of IL-6 and TNF-α as inflammatory cytokines. Furthermore, the level of G-CSF gene expression was significantly increased. In conclusion, M2000, a newly designed NSAID, has a remarkable effect on isolated PBMC in patients with MDS, which might bring a potential hope for its oral administrations in these patients.

Effects of guluronic acid (G2013) on gene expression of TLR2, TLR4, MyD88, TNF-α and CD52 in multiple sclerosis under in vitro conditions.

Context: Multiple sclerosis (MS) is an autoimmune and chronic inflammatory disease of CNS. The α-L-guluronic acid (G2013) as novel NSAID with immunomodulatory effects has shown its positive effects in various investigations.Objective: Present research aimed to study the potency of G2013 on gene expression of TLR2, TLR4, MyD88, TNF-α and CD52 in PBMCs of MS patients under in vitro conditions. Materials and methods: 24 blood samples from MS patients and healthy controls were considered for RT-PCR and flow cytometry techniques under two different doses of G2013.Results: Our research indicated that this drug could significantly decrease the gene expression of TLR2, TLR4 and TNF-α compared to untreated group. Conclusion: Data demonstrated that the guluronic acid is able to modify the expression levels of TLR2, TLR4 and TNF-α genes to less than the pathogenic boarder line level, which it might be recommended for reducing the pathological process in multiple sclerosis.

International multicenter randomized, placebo-controlled phase III clinical trial of ß-D-mannuronic acid in rheumatoid arthritis patients.

BACKGROUND: The oral administration of drug ß-D-mannuronic acid (M2000) showed a potent therapeutic effect in phase I/II study in rheumatoid arthritis (RA) patients. Here, our aim is to assess the efficacy and safety of this new drug in RA patients under a multinational, randomized placebo-controlled phase III clinical trial. METHOD: Patients (n = 288) with active disease at baseline and inadequate response to conventional drugs were randomly allocated to three groups; (1) receiving mannuronic acid at a dose of two capsules (500 mg) per day orally for 12 weeks, (2) placebo-controlled, and (3) conventional. The primary endpoints were the America College of Rheumatology 20 response (ACR20), 28-joint disease activity score (DAS28) and Modified Health Assessment Questionnaire-Disability Index (M-HAQ-DI). In addition, the participants were followed-up for safety assessment. RESULTS: In this phase III trial, after 12 weeks of treatment, there was a significant reduction in ACR20 between mannuronic-treated patients compared to placebo and conventional groups. Moreover, there was a similar significant improvement for DAS28 following mannuronic therapy. The statistical analysis showed a significant reduction in the swollen and tender joint count in mannuronic-treated patients compared with the placebo group. On the other side, mannuronic acid showed no-to-very low adverse events in comparison to placebo. CONCLUSION: The results of this multinational, phase III clinical trial provided a potent evidence base for the use of ß-D-mannuronic acid as a new highly safe and efficient drug in the treatment of RA.

Targeting of circulating Th17 cells by ß-D-mannuronic acid (M2000) as a novel medication in patients with rheumatoid arthritis.

OBJECTIVE: This study aimed at investigating the inhibitory effect of ß-D-mannuronic acid (M2000) on the Th17 circulating levels and IL-17 a related cytokine in rheumatoid arthritis (RA) patients. METHODS: The study included 27 patients with RA who had failed response to treatment. All patients were treated orally by M2000 at a dose of 500 mg twice daily for 12 weeks (Clinical trial identifier: IRCT2014011213739N2). The patients based on anti-tumor necrosis factor alpha (TNFα) blocker treatment were classified into two groups (conventional group and etanercept group). They were then allowed to continue their treatment excluding non-steroidal anti-inflammatory drugs (NSAIDs). The frequency of circulating Th17 cells and IL-17 serum level were determined before and 12 weeks after M2000 therapy and were compared to the healthy controls by using flow cytometry analysis and ELISA method, respectively. RESULTS: At baseline, higher circulating Th17 and IL-17 serum levels were significantly observed in both groups of RA patients than in the healthy controls (all P < 0.001). The frequency of Th17 cells significantly decreased in the conventional group as well as in the etanercept group after M2000 therapy but the level of reduction was higher in the conventional group compared to the etanercept group (P < 0.03 and P < 0.04, respectively). The IL-17 serum level significantly decreased in both groups after M2000 therapy (P < 0.01 and P < 0.02, respectively). Furthermore, the frequency of Th17 cells was positively correlated with Disease Activity Score (DAS28) (r = 0.34, P = 0.02). CONCLUSION: M2000 shows the inhibitory effect on the frequency of circulating Th17 cells as well as in the production of IL-17 in RA patients.

A phase I/II randomized, controlled, clinical trial for assessment of the efficacy and safety of ß-D-mannuronic acid in rheumatoid arthritis patients.

BACKGROUND: Following the potent efficacy of ß-D-mannuronic acid (M2000) in phase I/II trial in ankylosing spondylitis patients, the present clinical trial was conducted to evaluate the efficacy, safety, and tolerability of this novel drug in rheumatoid arthritis (RA) patients who had inadequate response to conventional therapy. METHOD: The study was a 12-week randomized, controlled, phase I/II clinical trial with two treatment arms: M2000 and conventional treatment. Patients who had RA according to the modified American College of Rheumatology (ACR) criteria, with active disease at baseline also inadequate response to conventional therapy, were enrolled in this study. M2000 was administrated at a dose of two capsules (500 mg) per day orally during a period of 12 weeks. The primary endpoint was the proportion of patients fulfilling the ACR 20% improvement criteria after 12 weeks of M2000 therapy. Moreover, the patients were also followed up for safety. RESULTS: There were no statistically significant differences between treatment and conventional groups at baseline characteristics. The ACR20 response rate was significantly higher among M2000-treated patients than conventional-treated control, so that 74% of patients in treatment group showed an ACR20 response after 12 weeks of M2000 therapy (74 versus 16%; P = 0.011). 10% of M2000-treated patients and 57.1% of conventional-treated patient's adverse events occurred during this study. CONCLUSION: Treatment with M2000 in combination with conventional therapy showed a significantly superior efficacy along with a high safety profile compared to conventional-treated patients. Thereby, M2000 might be suggested as a suitable option in the treatment of RA.

Therapeutic effects of pegylated-interferon-α2a in a mouse model of multiple sclerosis.

INTRODUCTION: Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis (MS). EAE is mainly mediated by adaptive and innate immune responses that lead to an inflammatory demyelination and axonal damage. The aim of the present research was to examine the therapeutic efficacy of Peg interferon alpha 2a (Peg-IFN α-2a) as a serine protease inhibitor on EAE model. MATERIAL AND METHODS: EAE induction was performed in female C57BL/6 mice by myelin oligodendrocyte glycoprotein (35-55) (MOG35-55) in Complete Freund's Adjuvant (CFA) emulsion, and Peg-IFN α-2a was used for the treatment of EAE. During the course of the study, clinical evaluation was assessed, and on day 21 post-immunisation blood samples were taken from the heart of mice for evaluation of IL-6, and enzymatic and non-enzymatic antioxidants. The mice were sacrificed and the brains and cerebellums were removed for histological analysis. RESULTS: Our findings indicated that Peg-IFN α-2a had beneficial effects on EAE by attenuation of the severity and a delay in the onset of disease. Histological analysis showed that treatment with Peg-IFN α-2a can reduce inflammation criteria. Moreover, in Peg-IFN α-2a-treated mice the serum level of IL-6 was significantly less than in controls, and total antioxidant capacity was significantly more than in the control animals. CONCLUSIONS: These data indicate that Peg-IFN α-2a as an anti-serine protease with immunomodulatory properties may be useful for the treatment of MS.

Preclinical and pharmacotoxicology evaluation of α-l-guluronic acid (G2013) as a non-steroidal anti-inflammatory drug with immunomodulatory property.

CONTEXT: Therapeutic effects of α-l-guluronic acid with the greatest tolerability and efficacy (G2013) have been shown in experimental model of multiple sclerosis and other in vitro and in vivo examinations regarding α-l-guluronic acid; there are no toxicological researches on its safety although the pharmacological impacts have been recorded. OBJECTIVE: This study was designed to determine the acute and sub chronic toxicity of α-l-guluronic acid in healthy male and female BALB/c mice. MATERIALS AND METHODS: For the acute toxicity study, the animals orally received five different single doses of α-l-guluronic acid and were kept under observation for 14 d. In the sub-chronic study, 24 male and female BALB/c mice were divided into four groups and treated daily with test substance preparation at dose levels of 0, 50, 250, and 1250 mg/kg body weight for at least 90 consecutive days. The mortality, body weight changes, clinical signs, hematological and biochemical parameters, gross findings, histopathological, and organs weight determinants were monitored during this study. RESULTS: The results of acute toxicity indicated that the LD50 of α-l-guluronic acid is 4.8 g/kg. We found no mortality or abnormality in clinical signs, body weight, relative organs weight, or necropsy in any of the animals in the subchronic study. Additionally, the results showed no significant difference in hematological, biochemical, and histopathological parameters in rats. CONCLUSIONS: Our results suggest that α-l-guluronic acid has high safety when administered orally in animals.

The reliability of clinical decisions based on the cervical vertebrae maturation staging method.

OBJECTIVES: Of the various techniques used to determine the optimum timing for growth modification treatments, the cervical vertebrae maturation method has great advantages, including validity and no need for extra X-ray exposure. Recently, the reproducibility of this method has been questioned. The aim of this study was to investigate the cause of poor reproducibility of this method and to assess the reproducibility of the clinical decisions made based on it. MATERIALS/METHODS: Seventy lateral cephalograms of Iranian patients aged 9‒15 years were observed twice by five experienced orthodontists. In addition to determining the developmental stage, each single parameter involved in this method was assessed in terms of inter- and intra-observer reproducibility. In order to evaluate the reproducibility of clinical decisions based on this method, cervical vertebrae maturation staging (CVMS) I and II were considered as phase 1 and CVMS IV and V were considered as phase 3. RESULTS: By considering the clinical approach of the CVMS method, inter-observer reproducibility of this method increased from 0.48 to 0.61 (moderate to substantial) and intra-observer reproducibility enhanced from 0.72 to 0.74. LIMITATIONS: 1. Complete visualization of the first four cervical vertebrae was an inclusion criterion, which also limits the clinical application of CVMS method. 2. These results can be generalized when determining growth modification treatments solely for Class II patients. CONCLUSIONS: Difficulty in determining the morphology of C3 and C4 leads to poor reproducibility of the CVMS method. Despite this, it has acceptable reproducibility in determining the timing of functional treatment for Class II patients.

Evaluation of therapeutic effects of nanofibrous mat containing mycophenolate mofetil on oral lichen planus: In vitro and clinical trial study.

Objectives: Recently, topical drug delivery system has gained increasing interest in the treatment of oral lesions. Lichen planus is a chronic inflammatory disease affecting mucous membranes and skin. The current study aimed to fabricate a drug delivery system containing mycophenolate mofetil for the treatment of oral lichen planus lesions. Methods: Firstly, a nanofibrous mat containing mycophenolate mofetil, zinc oxide nanoparticles, and aloe vera was designed and fabricated. The antimicrobial, cytocompatibility, anti-inflammatory, and antioxidative characteristics of fabricated scaffolds were evaluated. Then, this nanofibrous mat was applied to 12 patients suffering from bilateral erythematous/erosive Oral Lichen planus (OLP) lesions for 2 weeks. The treatment outcomes, including oral symptoms and lesion size, were compared with the routine topical treatment of these lesions; Triamcinolone ointment. Results: The characterization of nanofibrous mat approved the successful fabrication of scaffolds. The fabricated nanofibers showed notable antimicrobial activity. The amounts of TNF 𝛼, IL6, and reactive oxygen species (ROS) of stimulated human gingival fibroblasts were decreased after exposure to NFs/Myco/Alv/ZnO scaffolds. The clinical trial results demonstrated the same therapeutic effects compared to the commercial ointment, while the symptoms of patients were significantly improved in the mats group.Significance. Considering the successful results of this study, the application of nanofibrous mat can be a promising product for improving treatment outcomes of OLP.

The Open, Randomized, Positive Control Clinical Trial of Guluronic Acid (G2013) on SARS-CoV-2 Patients.

INTRODUCTION: Recently, the coronavirus disease 2019 (COVID-19) infection, with a vast spectrum of clinical and paraclinical symptoms has been a major health concern worldwide. Therapeutical management of COVID-19 includes antiviral and anti-inflammatory drugs. NSAIDs, as the second-line therapy, are often prescribed to relieve the symptoms of COVID-19. The α-L-guluronic acid (G2013) is a non-steroidal patented (PCT/EP2017/067920) agent with immunomodulatory properties. This study investigated the effect of G2013 on the outcome of COVID-19 in moderate to severe patients. METHODS: The disease's symptoms were followed up during hospitalization and for 4 weeks postdischarge in G2013 and control groups. Paraclinical indices were tested at the time of admission and discharge. Statistical analysis was performed on clinical and paraclinical parameters and ICU admission and death rate. RESULTS: The primary and secondary outcomes indicated the efficiency of G2013 on COVID-19 patients' management. There were significant differences in the duration of improvement of fever, coughing, fatigue/malaise. Also, a comparison of paraclinical indices at the time of admission and discharge showed significant change in prothrombin, D-dimer, and platelet. As the main findings of this study, G2013 significantly decreased the percentage of ICU admission (control:17 patients, G2013:1 patient) and death (control: 7 cases, G2013:0). CONCLUSION: These results conclude that G2013 has sufficient potential to be considered for moderate to severe COVID-19 patients, can significantly reduce the clinical and physical complications of this disease, has a positive effect on modulating the coagulopathy process, and aids in saving lives.

Adverse Effects of Anti-Covid-19 Drug Candidates and Alcohol on Cellular Stress Responses of Hepatocytes.

During the pandemic, dexamethasone (DEX), remdesivir (RDV), hydroxychloroquine (HCQ), thapsigargin (TG), camostat mesylate (CaM), and pralatrexate were repurposed drugs for coronavirus disease 2019 (COVID-19). However, the side effects on the liver associated with the anti-COVID therapies are unknown. Cellular stresses by these drugs at 0-30 µM were studied using HepG2, Huh7, and/or primary human hepatocytes. DEX or RDV induced endoplasmic reticulum stress with increased X-box binding protein 1 and autophagic response with increased accumulation of microtubule-associated protein 1A/1B-light chain 3 (LC3-II). DEX and RDV had additive effects on the stress responses in the liver cells, which further increased expression of activating transcription factor 4 and C/EBP homology protein 1 (CHOP), and cell death. Alcohol pretreatment (50 mM) and DEX induced greater cellular stress responses than DEX and RDV. Pralatrexate induced Golgi fragmentation, cell cycle arrest at G0/G1 phase, activations of poly (ADP-ribose) polymerase-1 (PARP) and caspases, and cell death. Pralatrexate and alcohol had synergistic effects on the cell death mediators of Bim, caspase3, and PARP. The protease inhibitor CaM and TG induced autophagic response and mitochondrial stress with altered mitochondrial membrane potential, B-cell lymphoma 2, and cytochrome C. TG and HCQ induced autophagic response markers of Unc-51 like autophagy activating kinase, LC3-II, Beclin1, and Atg5, and severe ER stress marker CHOP. Conclusion: These results suggest that the anti-COVID-19 drugs, especially with drug-drug or alcohol-drug combinations, cause cellular stress responses and injuries in the liver cells.

Novel nanocomposite scaffold based on gelatin/PLGA-PEG-PLGA hydrogels embedded with TGF-ß1 for chondrogenic differentiation of human dental pulp stem cells in vitro.

In the current study, a novel nanocomposite hydrogel scaffold comprising of natural-based gelatin and synthetic-based (poly D, L (lactide-co-glycolide) -b- poly (ethylene glycol)-b- poly D, L (lactide-co-glycolide) (PLGA-PEG-PLGA) triblock copolymer was developed and loaded with transforming growth factor- ß1 (TGF-ß1). Synthesized scaffolds' chemical structure was examined by 1H NMR and ATR-FTIR. Scanning electron microscopy (SEM) confirmed particle size and morphology of the prepared nanoparticles as well as the scaffolds. The morphology analysis revealed a porous interconnected structure throughout the scaffold with a pore size dimension of about 202.05 µm. The swelling behavior, in vitro degradation, mechanical properties, density, and porosity were also evaluated. Phalloidin/DAPI staining was utilized for confirming the extended cytoskeleton of the chondrocytes. Alcian blue staining was conducted to determine cartilaginous matrix sulfated glycosaminoglycan (sGAG) synthesis. Eventually, over a period of 21 days, a real-time RT-PCR analysis was applied to measure the mRNA expression of chondrogenic marker genes, type-II collagen, SOX 9, and aggrecan, in hDPSCs cultured for up to 21 days to study the influence of gelatin/PLGA-PEG-PLGA-TGF-ß1 hydrogels on hDPSCs. The findings of the cell-encapsulating hydrogels analysis suggested that the adhesion, viability, and chondrogenic differentiation of hDPSCs improved by gelatin/PLGA-PEG-PLGA-TGF-ß1 nanocomposite hydrogels. These data supported the conclusion that gelatin/PLGA-PEG-PLGA-TGF-ß1 nanocomposite hydrogels render the features that allow thein vitrofunctionality of encapsulated hDPSCs and hence can contribute the basis for new effective strategies for the treatment of cartilage injuries.

MTA-Enriched Polymeric Scaffolds Enhanced the Expression of Angiogenic Markers in Human Dental Pulp Stem Cells.

Revascularization of the pulp tissue is one of the fundamental processes and challenges in regenerative endodontic procedures (REPs). In this regard, the current study is aimed at synthesizing the mineral trioxide aggregate- (MTA-) based scaffolds as a biomaterial for REPs. Poly (ε-caprolactone) (PCL)/chitosan (CS)/MTA scaffolds were constructed and evaluated by FTIR, SEM, XRD, and TGA analyses. Proliferation and adhesion of human dental pulp stem cells (hDPSCs) were assessed on these scaffolds by scanning electron microscopy (SEM) and MTT assays, respectively. The expression of angiogenic markers was investigated in gene and protein levels by real-time PCR and western blotting tests. Our results indicated that the obtained appropriate physicochemical characteristics of scaffolds could be suitable for REPs. The adhesion and proliferation level of hDPSCs were significantly increased after seeding on PCL/CS/MTA scaffolds. The expression levels of VEGFR-2, Tie2, and Angiopoietin-1 genes were statistically increased on the PCL/CS/MTA scaffold. In support of these findings, western blotting results showed the upregulation of these markers at protein levels in PCL/CS/MTA scaffold (P < 0.05). The current study results suggested that PCL/CS/MTA scaffolds provide appropriate structures for the adhesion and proliferation of hDPSCs besides induction of the angiogenesis process in these cells.

Enhancing the function of PLGA-collagen scaffold by incorporating TGF-ß1-loaded PLGA-PEG-PLGA nanoparticles for cartilage tissue engineering using human dental pulp stem cells.

Since cartilage has a limited capacity for self-regeneration, treating cartilage degenerative disorders is a long-standing difficulty in orthopedic medicine. Researchers have scrutinized cartilage tissue regeneration to handle the deficiency of cartilage restoration capacity. This investigation proposed to compose an innovative nanocomposite biomaterial that enhances growth factor delivery to the injured cartilage site. Here, we describe the design and development of the biocompatible poly(lactide-co-glycolide) acid-collagen/poly(lactide-co-glycolide)-poly(ethylene glycol)-poly(lactide-co-glycolide) (PLGA-collagen/PLGA-PEG-PLGA) nanocomposite hydrogel containing transforming growth factor-ß1 (TGF-ß1). PLGA-PEG-PLGA nanoparticles were employed as a delivery system embedding TGF-ß1 as an articular cartilage repair therapeutic agent. This study evaluates various physicochemical aspects of fabricated scaffolds by 1HNMR, FT-IR, SEM, BET, and DLS methods. The physicochemical features of the developed scaffolds, including porosity, density, degradation, swelling ratio, mechanical properties, morphologies, BET, ELISA, and cytotoxicity were assessed. The cell viability was investigated with the MTT test. Chondrogenic differentiation was assessed via Alcian blue staining and RT-PCR. In real-time PCR testing, the expression of Sox-9, collagen type II, and aggrecan genes was monitored. According to the results, human dental pulp stem cells (hDPSCs) exhibited high adhesion, proliferation, and differentiation on PLGA-collagen/PLGA-PEG-PLGA-TGFß1 nanocomposite scaffolds compared to the control groups. SEM images displayed suitable cell adhesion and distribution of hDPSCs throughout the scaffolds. RT-PCR assay data displayed that TGF-ß1 loaded PLGA-PEG-PLGA nanoparticles puts forward chondroblast differentiation in hDPSCs through the expression of chondrogenic genes. The findings revealed that PLGA-collagen/PLGA-PEG-PLGA-TGF-ß1 nanocomposite hydrogel can be utilized as a supportive platform to support hDPSCs differentiation by implementing specific physio-chemical features.