RJX Improves Wound Healing in Diabetic Rats.

Background: We recently reported the clinical safety profile of RJX, a well-defined intravenous GMP-grade pharmaceutical formulation of anti-oxidant and anti-inflammatory vitamins as active ingredients, in a Phase 1 study in healthy volunteers (ClinicalTrials.gov Identifier: NCT03680105) (Uckun et al., Front. Pharmacol. 11, 594321. 10.3389/fphar.2020.594321). The primary objective of the present study was to examine the effects of GMP-grade RJX on wound and burn injury healing in diabetic rats. Methods: In the present study, a rat model of T2DM was used that employs HFD in combination with a single injection of STZ intraperitoneally (i.p) at a moderate dose level (45 mg/kg). Anesthetized diabetic rats underwent full-thickness skin excision on the back or were subjected to burn injury via a heated brass probe and then started on treatments with normal saline (NS = vehicle) or RJX administered via intraperitoneal injections for three weeks. Findings: Notably, diabetic rats treated with the 1.25 mL/kg or 2.5 mL/kg RJX (DM+RJX groups) rapidly healed their wounds as fast as non-diabetic control rats. Inflammatory cell infiltration in the dermis along with fibrin and cell debris on the epithelial layer persisted for up to 14 days in the DM+NS group but not in RJX-treated groups. The histopathological score of wound healing on days 7 and 14 was better in diabetic rats treated with RJX than diabetic rats treated with NS and comparable to the scores for non-diabetic healthy rats consistent with an accelerated healing process. The residual wound area of RJX-treated rats was significantly smaller than that of NS-treated diabetic rats at each evaluation time point (P<0.001). The accelerating effect of RJX on diabetic wound healing was dose-dependent. We obtained similar results in the burn injury model. Our results demonstrate that RJX - at a dose level >10-fold lower than its clinical maximum tolerated dose (MTD) - accelerates the healing of excision wounds as well burn injury in diabetic rats.

Protective effect of a novel polyherbal formulation on experimentally induced osteoarthritis in a rat model.

Osteoarthritis (OA) is a musculoskeletal disorder mainly found in elderly individuals. Modern treatment of OA, like nonsteroidal anti-inflammatory drugs, corticosteroids, hyaluronic acid injections, etc., is linked to long-term side effects. We evaluated the anti-osteoarthritic properties of a novel joint health formula (JHF) containing Bisdemethoxycurcumin enriched curcumin, 3-O-Acetyl-11-keto-beta-Boswellic acid-enriched Boswellia, and Ashwagandha in monosodium iodoacetate (MIA)-induced knee OA in rats. Twenty-eight female rats were distributed into four groups: Control, OA, OA + JHF (100 mg/kg), and OA + JHF (200 mg/kg). JHF decreased the right joint diameters but increased the paw area and stride length compared to the OA group with no treatment. JHF significantly reduced the arthritic conditions after four weeks of supplementation (p < 0.05). JHF significantly decreased TNF-α, IL-1ß, IL-10, COMP, and CRP in the serum of osteoarthritic rats (p < 0.0001). We observed reduced lipid peroxidation but increased SOD, GSH-Px, and CAT activities in response to JHF treatment in OA animals. JHF down-regulated MMP-3, COX-2, and LOX-5 and improved the histological structure of the knee joint of osteoarthritic rats. JHF demonstrated a protective effect against osteoarthritis, possibly due to anti-inflammatory and antioxidant activity in experimentally induced osteoarthritis in rats, and could be an effective option in the management of OA.

Niacinamide and undenatured type II collagen modulates the inflammatory response in rats with monoiodoacetate-induced osteoarthritis.

The current work aimed to examine the properties of oral supplementation of niacinamide and undenatured type II collagen (UCII) on the inflammation and joint pain behavior of rats with osteoarthritis (OA). Forty-nine Wistar rats were allocated into seven groups; control (no MIA), MIA as a non-supplemental group with monosodium iodoacetate (MIA)-induced knee osteoarthritis, MIA + undenatured type II collagen (UCII) at 4 mg/kg BW, MIA + Niacinamide at 40 mg/kg BW (NA40), MIA + Niacinamide at 200 mg/kg BW (NA200), MIA + UCII + NA40 and MIA + UCII + NA200. Serum IL-1ß, IL-6, TNF-α, COMP, and CRP increased in rats with OA and decreased in UCII and NA groups (p < 0.05). Rats with osteoarthritis had greater serum MDA and knee joint MMP-3, NF-κB, and TGß protein levels and decreased in treated groups with UCII and NA (p < 0.05). The rats with OA also bore elevated joint diameters with joint pain behavior measured as decreased the stride lengths, the paw areas, and the paw widths, and increased the Kellgren-Lawrence and the Mankin scores (p < 0.05) and decreased in UCII treated groups. These results suggest the combinations with the UCII + NA supplementation as being most effective and reduce the inflammation responses for most OA symptoms in rats.

A Next Generation Formulation of Curcumin Ameliorates Experimentally Induced Osteoarthritis in Rats via Regulation of Inflammatory Mediators.

Osteoarthritis (OA) is a chronic and debilitating disease of the knee joint. OA of the knee is initiated by physical damage and accumulated oxidative stress, followed by an exaggerated inflammation leading to cartilage damage. Currently, no effective and safe therapeutic option capable of restoring articular cartilage tissue and joint architecture is available. We here report a novel and highly bioavailable formulation of curcumin, labeled as Next Generation Ultrasol Curcumin (NGUC), which was 64.7 times more bioavailable than natural 95% curcumin extract as demonstrated in rat bioavailability studies. We further investigated the protective effect of NGUC against monosodium iodoacetate (MIA)-induced knee OA in rats. Analysis of X-ray and histopathological images revealed that NGUC supplementation restored joint architecture and reduced swelling of joints induced by MIA. NGUC treatment caused a significant reduction in the levels of inflammatory mediators such as TNF-α, IL-1ß, IL-6, COMP, and CRP, and expressions of MMP-3, 5-LOX, COX-2, and NFκB in synovial tissue of rats with MIA-induced OA. NGUC also decreased serum MDA level and increased the levels of antioxidant enzymes SOD, CAT, and GPX. Thus, our results indicate that a novel formulation of curcumin with enhanced bioavailability effectively ameliorates the pathophysiology of OA.

Undenatured Type II Collagen Ameliorates Inflammatory Responses and Articular Cartilage Damage in the Rat Model of Osteoarthritis.

Osteoarthritis (OA) is an age-related joint disease that includes gradual disruption of the articular cartilage and the resulting pain. The present study was designed to test the effects of undenatured type II collagen (UC-II®) on joint inflammation in the monoiodoacetate (MIA) OA model. We also investigated possible mechanisms underlying these effects. Female Wistar rats were divided into three groups: (i) Control; (ii) MIA-induced rats treated with vehicle; (iii) MIA-induced rats treated with UC-II (4 mg/kg BW). OA was induced in rats by intra-articular injection of MIA (1 mg) after seven days of UC-II treatment. UC-II reduced MIA-induced Kellgren-Lawrence scoring (53.3%, P < 0.05). The serum levels of inflammatory cytokines [IL-1ß (7.8%), IL-6 (18.0%), TNF-α (25.9%), COMP (16.4%), CRP (32.4%)] were reduced in UC-II supplemented group (P < 0.0001). In the articular cartilage, UC-II inhibited the production of PGE2 (19.6%) and the expression of IL-1ß, IL-6, TNF-a, COX-2, MCP-1, NF-κB, MMP-3, RANKL (P < 0.001). The COL-1 and OPG levels were increased, and MDA decreased in UC-II supplemented rats (P < 0.001). UC-II could be useful to alleviate joint inflammation and pain in OA joints by reducing the expression of inflammatory mediators.

Arginine Silicate Inositol Complex Accelerates Cutaneous Wound Healing.

Arginine silicate inositol (ASI) complex is a composition of arginine, silicon, and inositol that has been shown to have beneficial effects on vascular health. This study reports the effects of an ASI ointment on wound healing in rats. A full-thickness excision wound was created by using a disposable 5 mm diameter skin punch biopsy tool. In this placebo-controlled study, the treatment group's wound areas were covered by 4 or 10 % ASI ointments twice a day for 5, 10, or 15 days. The rats were sacrificed either 5, 10, or 15 days after the wounds were created, and biopsy samples were taken for biochemical and histopathological analysis. Granulation tissue appeared significantly faster in the ASI-treated groups than in the control groups (P < 0.05). The mean unhealed wound area was significantly smaller, and the mean percentage of total wound healing was significantly higher in ASI-treated wounds than in the control wounds. Hydroxyproline, collagen, and matrix metalloproteinases were measured in the granulated tissue and found to be affected. Inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), collagen, matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and various cytokines (TNF-α and IL-1ß) measured in this study showed a significant fall in expression level in ASI-treated wounds. The results suggest that topical application of ASI ointment (especially 4 % concentration) has beneficial effects on the healing response of an excisional wound.

Effects of sildenafil citrate on torsion/detorsion-induced changes in red blood cell and plasma lipid peroxidation, antioxidants, and blood hematology of male rats.

OBJECTIVE: The aim of this work was to determine effects of intrapertoneally-administered sildenafil citrate (SC) for prevention testicular injury after unilateral testicular torsion/detorsion (T/D) in rats on red blood cell (RBC) and plasma lipid peroxidation, antioxidants and blood hematology. STUDY DESIGN: Thirty seven adult male Wistar albino rats were divided into four groups: sham operated (group 1), T/D+saline (group 2), T/D+0.7mg SC (group 3) and T/D+1.4mg SC (group 4). Testicular torsion was created by rotating the right testis 720° in a clockwise direction for 2h in all the groups, except for group 1. RESULTS: Our results showed that that testicular injury significantly induced erythrocyte reduced glutathion (GSH) (p<0.05), malondialdehyde (MDA) in RBC (p<0.01) and plasma (p<0.05) and blood lymphocyte (p<0.01) counts. Administration of low dose SC led to significantly increase in the levels of RBC GSH (p<0.05), plasma paraoxonase (PON1) (p<0.01), nitric oxide (NO) (p<0.01) and blood lymphocyte counts (p<0.01), but to decreases in the levels of MDA in plasma and RBC, blood mean corpuscular volume (MCV) (p<0.05) and eosinophil counts (p<0.05). Treatment with high dose SC caused a significantly increase in PON1, vitamin E and ß-carotene in plasma, levels of GSH in RBC and blood lymphocyte counts. On the other hand, results showed that high dose sildenafil significantly decreased plasma and RBC MDA levels. Total tissue damage scores of the group 2 were significantly higher than group 1 and 3. CONCLUSION: Low dose SC appears to be beneficial in reducing the effects of injury to the testicular torsion.

Efficacy of vitamin E and selenium for the prevention of intra-abdominal adhesions in rats: uterine horn models.

OBJECTIVE: This study compares the efficacies of vitamin E and selenium, both individually and in combination, for the prevention of postoperative intra-abdominal adhesions in rats. METHODS: Forty-seven female rats were divided into five groups. The sham animals (S group, n = 7) were given only laparotomies and intraperitoneally received 0.9% NaCl (2 ml). In the 40 other rats, abrasions of the left uterine horn were performed, followed by intraperitoneal administration of either 2 ml 0.9% NaCl (C group), 10 mg vitamin E (vitamin E group), 0.2 mg/kg selenium (Se group) or 10 mg vitamin E with 0.2 mg/kg selenium (vitamin E + Se group), with 10 animals in each treatment group. RESULTS: Adhesion formation was significantly reduced in animals in the Se and vitamin E + Se groups (p<0.05). Tissue catalase and glutathione peroxidase activities did not significantly differ between the groups. However, catalase and glutathione peroxidase activities and reduced glutathione levels were slightly increased in the vitamin E, Se and vitamin E + Se groups. In the vitamin E group, malondialdehyde concentrations were significantly lower than in the C group (p<0.05), but no significant differences were present among the S, C, Se and vitamin E + Se groups. Levels of nitric oxide were significantly higher in the C group than in the other groups (p<0.01). CONCLUSION: Intraperitoneal administration of selenium or combined vitamin E and selenium appears to be effective in preventing intra-abdominal adhesion formation in rat models through the reduction of lipid peroxidation products.

Protective effect of sildenafil citrate on contralateral testis injury after unilateral testicular torsion/detorsion.

OBJECTIVES: This study was designed to investigate prevention of contralateral testicular injury with sildenafil citrate after unilateral testicular torsion/detorsion. METHODS: Thirty-seven adult male rats were divided into four groups: sham operated (group 1, n = 7), torsion/detorsion + saline (group 2, n = 10), torsion/detorsion + 0.7 mg of sildenafil citrate (group 3, n = 10) and torsion/detorsion + 1.4 mg of sildenafil citrate (group 4, n = 10). Unilateral testicular torsion was created by rotating the right testis 720º in a clockwise direction for 2 h in other groups, except for group 1, which was served as sham group. After torsion (2 h) and detorsion (2 h) periods, rats were killed. RESULTS: The level of reduced glutathion (GSH) (p < 0.05) and the activities of catalase (p < 0.01) and glutathione peroxidase (p < 0.05) in the contralateral testis from group 2 were significantly lower and nitric oxide (NO) (p < 0.05) level in the contralateral testis were significantly higher than those of group 1. Administration of low-dose sildenafil citrate (group 3) prevented the increases in malondialdehyde and NO levels and decreases in glutathione peroxidase activities and GSH values induced by testicular torsion. However, administration of high-dose sildenafil citrate (group 4) had no effect on these testicular parameters (p > 0.05). Histopathological changes were detected in groups 2, 3 and 4. CONCLUSION: These results suggest that biochemically and histologically torsion/detorsion injury occurs in the contralateral testis following 2-h torsion and 2-h detorsion and that administration of low-dose sildenafil citrate before detorsion prevents ischemia/reperfusion cellular damage in testicular tissue.

The comparison of methylene blue and vitamin E in prevention of abdominal postoperative adhesion formation in rat uterine horn models: Biochemical and histopathologic evaluation / Comparação do azul de metileno e vitamina E na prevenção de aderência abdominal pós-operatória em corno uterino de ratos: avalição bioquímica e histopatológica

Purpose: To compare the effects of vitamin E and 1 percent methylen blue solutions on prevention of experimentally induced adhesions in rats. Methods: Thirty seven female Spraque Dawley rats were randomized into four groups. First group was kept as sham operated group. An adhesion model was constituted on the left uterine horn of the other groups. The lesion areas of rats from the second, the third and the fourth groups were coated with 2 ml 0.9 percent saline solution (C group), 10 mg vitamin E (VE group) and 1 percent methylen blue solutions (MB group), respectively. Results: Histopathologically, adhesion scores, mononuclear cell infiltration, oedema and fibrosis were more prominent in the MB group compared with C and VE groups. There were no significant differences between the groups in tissue glutathione peroxidase (GPx), catalase (CAT) activities and glutation (GSH) level, these parameters were slightly increased in group with VE supplementation though. The administration of VE and MB significantly decreased NO (P<0.01) levels when compared to the C group. The level of malondialdehyde (MDA) in the VE group was significantly lower (P<0.05) than those of the Sh and C groups. Conclusion: Intraperitoneal methylen blue solutions treatments were more effective according to vitamin E in preventing the formation of intra-abdominal adhesion in a rat uterine horn model.

Objetivo: Comparar os efeitos da vitamina E e 1 por cento da solução de azul de metileno na prevenção de aderências induzidas em ratos. Métodos: Trinta e sete ratos fêmeas Spraque Dawley foram distribuídos em quatro grupos. O primeiro grupo foi mantido como grupo sham. O modelo de aderência foi realizado no corno uterino esquerdo nos outros grupos. As áreas da lesão dos ratos do segundo, terceiro e quarto grupos foram revestidas com 2 ml de solução salina 0,9 por cento (Grupo C), 10 mg de vitamina E (Grupo VE) e solução de azul de metileno 1 por cento (Grupo MB), respectivamente. Resultados: Histopatologicamente, o escore das aderências, infiltração celular mononuclear, edema e fibrose foram mais proeminentes no grupo MB em comparação aos grupos C e VE. Não houve diferença significante entre os grupos na peroxidase da glutatione do tecido (GPx), atividade da catalase (CAT) e o nível de glutation (GSH). Estes parâmetros foram ligeiramente aumentados no grupo com suplemento da VE. A administração da VE e do MB diminuiu significantemente os níveis quando quando comparada ao Grupo C. O nível de malondialdeído no grupo VE foi significantemente mais baixo do que nos grupos sham e C. Conclusão: A administração intraperitoneal da solução de azul de metileno foi mais eficaz de acordo com a vitamina E na prevenção de aderências intra-abdominais no corno uterino de ratos.

The comparison of methylene blue and vitamin E in prevention of abdominal postoperative adhesion formation in rat uterine horn models: Biochemical and histopathologic evaluation.

PURPOSE: To compare the effects of vitamin E and 1% methylen blue solutions on prevention of experimentally induced adhesions in rats. METHODS: Thirty seven female Spraque Dawley rats were randomized into four groups. First group was kept as sham operated group. An adhesion model was constituted on the left uterine horn of the other groups. The lesion areas of rats from the second, the third and the fourth groups were coated with 2 ml 0.9 % saline solution (C group), 10 mg vitamin E (VE group) and 1% methylen blue solutions (MB group), respectively. RESULTS: Histopathologically, adhesion scores, mononuclear cell infiltration, oedema and fibrosis were more prominent in the MB group compared with C and VE groups. There were no significant differences between the groups in tissue glutathione peroxidase (GPx), catalase (CAT) activities and glutation (GSH) level, these parameters were slightly increased in group with VE supplementation though. The administration of VE and MB significantly decreased NO (P<0.01) levels when compared to the C group. The level of malondialdehyde (MDA) in the VE group was significantly lower (P<0.05) than those of the Sh and C groups. CONCLUSION: Intraperitoneal methylen blue solutions treatments were more effective according to vitamin E in preventing the formation of intra-abdominal adhesion in a rat uterine horn model.

Surgery-induced changes in red blood cell and plasma lipid peroxidation, enzymatic and non-enzymatic antioxidants, and blood hematology of female rats: protective role of methylene blue and vitamin E.

OBJECTIVE: The aim was to determine the effects on red blood cell (RBC) and plasma lipid peroxidation, antioxidants and blood hematology of intraperitoneally administered vitamin E (VE) and 1% methylene blue (MB) solutions for prevention of adhesions in rats. STUDY DESIGN: Thirty-seven female Sprague-Dawley rats were randomized into four groups. An adhesion model was constituted on the left uterine horn in three of the groups. They were then given intraperitoneally 0.9% saline (C group), 10 mg VE (VE group) and 1% MB (MB group) solutions, respectively. A sham group (Sh group), on which laparotomy was performed, received 2 ml of 0.9% saline solution. RESULTS: In the C group, the adhesion scores were significantly higher than in the VE (p<0.01), MB (p<0.01) and Sh groups (p<0.005). Results showed that adhesion formation significantly induced nitric oxide (NO) (p<0.01) and malondialdehyde (MDA) in plasma (p<0.01). The levels of RBC glutathione (GSH) (p<0.05) and plasma VE (p<0.01) significantly increased after VE administration, whereas the levels of MDA (RBC and plasma) (p<0.01), plasma NO (p<0.01), blood lymphocyte count (p<0.05) and blood white blood cell (WBC) counts (p<0.01) decreased. Treatment with MB caused a significant increase in plasma VE (p<0.01). On the other hand, results showed that MB significantly decreased blood WBC counts (p<0.01), plasma paraoxonase (PON1) (p<0.001) and NO (p<0.01), and RBC glutathione peroxidase (GPx) activity (p<0.05) and MDA levels (p<0.01). CONCLUSION: Intraperitoneal administration of MB and VE is significantly effective in preventing intraabdominal adhesion formation in a rat model. Further investigations are necessary, however, to better understand the underlying biochemical mechanisms on lipid peroxidation and antioxidants of MB.

Protective effect of sildenafil citrate on contralateral testis injury after unilateral testicular torsion/detorsion

OBJECTIVES: This study was designed to investigate prevention of contralateral testicular injury with sildenafil citrate after unilateral testicular torsion/detorsion. METHODS: Thirty-seven adult male rats were divided into four groups: sham operated (group 1, n = 7), torsion/detorsion + saline (group 2, n = 10), torsion/detorsion + 0.7 mg of sildenafil citrate (group 3, n = 10) and torsion/detorsion + 1.4 mg of sildenafil citrate (group 4, n = 10). Unilateral testicular torsion was created by rotating the right testis 720º in a clockwise direction for 2 h in other groups, except for group 1, which was served as sham group. After torsion (2 h) and detorsion (2 h) periods, rats were killed. RESULTS: The level of reduced glutathion (GSH) (p<0.05) and the activities of catalase (p<0.01) and glutathione peroxidase (p<0.05) in the contralateral testis from group 2 were significantly lower and nitric oxide (NO) (p<0.05) level in the contralateral testis were significantly higher than those of group 1. Administration of low-dose sildenafil citrate (group 3) prevented the increases in malondialdehyde and NO levels and decreases in glutathione peroxidase activities and GSH values induced by testicular torsion. However, administration of high-dose sildenafil citrate (group 4) had no effect on these testicular parameters (p>0.05). Histopathological changes were detected in groups 2, 3 and 4. CONCLUSION: These results suggest that biochemically and histologically torsion/detorsion injury occurs in the contralateral testis following 2-h torsion and 2-h detorsion and that administration of low-dose sildenafil citrate before detorsion prevents ischemia/reperfusion cellular damage in testicular tissue.

Efficacy of vitamin E and selenium for the prevention of intra-abdominal adhesions in rats: uterine horn models

OBJECTIVE: This study compares the efficacies of vitamin E and selenium, both individually and in combination, for the prevention of postoperative intra-abdominal adhesions in rats. METHODS: Forty-seven female rats were divided into five groups. The sham animals (S group, n = 7) were given only laparotomies and intraperitoneally received 0.9 percent NaCl (2 ml). In the 40 other rats, abrasions of the left uterine horn were performed, followed by intraperitoneal administration of either 2 ml 0.9 percent NaCl (C group), 10 mg vitamin E (vitamin E group), 0.2 mg/kg selenium (Se group) or 10 mg vitamin E with 0.2 mg/kg selenium (vitamin E + Se group), with 10 animals in each treatment group. RESULTS: Adhesion formation was significantly reduced in animals in the Se and vitamin E + Se groups (p<0.05). Tissue catalase and glutathione peroxidase activities did not significantly differ between the groups. However, catalase and glutathione peroxidase activities and reduced glutathione levels were slightly increased in the vitamin E, Se and vitamin E + Se groups. In the vitamin E group, malondialdehyde concentrations were significantly lower than in the C group (p<0.05), but no significant differences were present among the S, C, Se and vitamin E + Se groups. Levels of nitric oxide were significantly higher in the C group than in the other groups (p<0.01). CONCLUSION: Intraperitoneal administration of selenium or combined vitamin E and selenium appears to be effective in preventing intra-abdominal adhesion formation in rat models through the reduction of lipid peroxidation products.