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Background: The human amniotic membrane (HAM) is a suitable and effective scaffold for cell culture and delivery, and adipose-derived stem cells (ADSCs) are an important source of stem cells for transplantation and chondrogenic differentiation. Objective: To assess the practicability of a cryopreserved HAM as a scaffold in cell proliferation and differentiation in vitro. Materials and Methods: In this experimental study, adipose tissue samples were harvested from the inguinal region of male patients aged 15-30 years. Flow cytometry was used to identify CD31, CD45, CD90, and CD105 markers in adipose stem cells. HAM was harvested from donor placenta after cesarean section, washed, trypsin-based decellularized trypsinized decellularized, and used as a scaffold via three methods: 1) ADSCs were differentiated into chondrocytes on cell culture flasks (monolayer method), and after 14 days of culture, the cells were transferred and cultured on both sides of the HAM; 2) ADSCs were cultured and differentiated directly on both sides of the HAM for 14 days (scaffold-mediated differentiation); and 3) chondrocytes were differentiated with micromass culture for 14 days, transferred on HAM, and tissue slides were histologically analyzed qualitatively. Results: Flow cytometry confirmed the presence of mesenchymal stem cells. Histological findings revealed that the cells adhered and grew well on the stromal layer of HAM. Among the three methods, scaffold-mediated differentiation of ADSCs showed the best results. Conclusion: ADSCs have excellent attachment, viability, and differentiation capacity in the stromal side of HAM. Additionally, the direct culture and differentiation of ADSCs on HAM is more suitable than the culture of differentiated cells on HAM. Key words: Amniotic membrane, Scaffold, Chondrogenesis, Differentiation, Mesenchymal stem cell.

Human amniotic membrane (HAM) is traditionally used for the treatment of non-healing wounds. However, high density of HAM-matrix (HAM-M) diminishes cellular contribution for successful tissue regeneration. Herein we investigated whether a bioengineered micro-porous three-dimensional (3D) HAM-scaffold (HAM-S) could promote healing in ischemic wounds in diabetic type 1 rat. HAM-S was prepared from freshly decellularized HAM. Then, 30 days after inducing diabetes, an ischemic circular excision was generated on rats’ skin. The diabetic animals were randomly divided into untreated (Diabetic group), engrafted with HAM-M (D-HAM-M group) and HAM-S (D-HAM-S group). Also, non-diabeticuntreated rats (Healthy group) were considered as control. Stereological, molecular, and tensiometrical assessments were performed on post-surgical days 7, 14, and 21. We found that the volumes of new epidermis and dermis, the numerical density of epidermal basal cells and fibroblasts, the length density of blood vessels, the numbers of proliferating cells and collagen deposition as well as biomechanical properties of healed wound were significantly higher in D-HAM-S group in most cases compared those of the diabetic group, or even in some cases compared to D-HAM-M group. Furthermore, in D-HAM-S group, the transcripts for genes contributing to regeneration ( Tgf-β , bFgf and Vegf ) upregulated more than those of D-HAM-M group, when compared to diabetic ones. Overall, the HAM-S had more impact on delayed wound healing process compared to traditional use of intact HAM. It is therefore suggested that the bioengineered three dimensional micro-porous HAM-S is more suitable for cells adhesion, penetration, and migration for contributing to wounded tissue regeneration.

Background Damage to peripheral nerves results in sensory, motor, and autonomic impairments, which are frequently accompanied by neuropathic pain. Effective strategies for nerve regeneration remain a major clinical challenge. Methods We engineered nanofibrous nerve guidance conduits (NGCs) composed of poly(ε-caprolactone) (PCL), multiwalled carbon nanotubes (CNTs), and epigallocatechin gallate (EGCG), which were fabricated by electrospinning and subsequently filled with a collagen hydrogel. CNTs were incorporated to improve the mechanical strength, physicochemical properties, and electrical conductivity, whereas EGCG had anti-inflammatory and antioxidant effects. The scaffold morphology was evaluated by scanning electron microscopy (SEM) and atomic force microscopy (AFM). In vitro assays were used to assess mesenchymal stem cell (MSC) viability and morphology. Functional recovery was examined via sciatic nerve functional indices and gastrocnemius electromyography. Results At 12 weeks post-axotomy, stereological, immunohistochemical, MRI, and real-time PCR analyses of sciatic nerves and L4–L5 dorsal root ganglia revealed significant upregulation of neuronal markers (MAP2, β-tubulin III, and neurofilament) and Schwann cell markers (S100, and NF-200), suggesting enhanced neuronal maturation and myelination. Compared with the control conditions, the composite conduits promoted improved motor function, nerve conduction velocity, and muscle preservation. Conclusion PCL/CNT/EGCG nanofibrous conduits, combined with a collagen hydrogel provide a favorable microenvironment for peripheral nerve regeneration. This strategy has translational potential for improving outcomes following peripheral nerve injury. Graphical abstract

Background Diabetic wounds are one of the most important issues in diabetic patients. It seems that Juglans regia L. leaf with antioxidant and anti-inflammatory potentials can be profitable for healing of diabetic wounds. The aim of present study was to investigate the topical administration of Juglans regia L. leaf extract in diabetic wound healing. Methods Seventy-five diabetic male rats were randomly divided into 5 groups ( n  = 15), including: untreated (Control) group, Eucerin group, 2% Juglans regia L . ointment (JRL 2%) group, 5% Juglans regia L . ointment (JRL 5%) group, and Phenytoin group as a reference drug. Sampling was performed at days 7, 14, and 21 after surgery. Evaluation tests included stereology, immunohistochemistry, molecular, and biomechanical. Results Our results showed that the wound closure rate, volumes of newly formed of epidermis and dermis, density of fibroblasts and blood vessels, collagen deposition, density of proliferation cells, expression levels of TGF-β and VEGF genes, and biomechanical characteristics were significantly higher in extract groups compared to control and eucerin groups, however, these changes were considerable in the JRL 5% group ( P  <  0.05 ). This is while that the density of neutrophils and expression levels of TNF-α and IL-1β genes in the extract groups, especially in the JRL 5% group, were significantly reduced compared to control and eucerin groups ( P  <  0.05 ). Conclusion Topical administration of Juglans regia L. leaf extract, especially in 5% concentration, considerably accelerates diabetic wound healing.

Background: Cyclophosphamide (CP), as a chemotherapy drug, causes severe damage in testicular tissue through producing free radicals. Cerium oxide nanoparticles (NC) exhibit antioxidant and anti-inflammatory properties. The purpose of this study was to investigate the protective effect of NC on CP-induced testicular damage in mice. Methods: In this experimental study, thirty-two male mice were divided into four groups (eight mice in each group). The control group was received intraperitoneally (IP) normal saline, NC group was received NC for three consecutive days (100 μg/kg, IP), CP group was received CP (200 mg/kg, IP), and the CP + NC group received NC, three consecutive days before receiving CP. After 2 days, testicles were assessed for biochemical, histomorphometrical, histopathological, and immunohistochemical analyses. Results: CP administration caused statistically significant increases in sperm abnormality, malondialdehyde, protein carbonyl levels, reactive oxygen species, level and apoptosis, and decreases in sperm count, sperm viability, testosterone, glutathione activity, the mean thickness of the germinal epithelium, diameter of seminiferous tubules in mice. Degeneration, necrosis, arrest of spermatogenesis, congestion, and atrophy in testicular tissue confirmed the low Johnsen's Testicular score in CP group. Administration of NC significantly ameliorated the CP-induced adverse effects on testis compared with the CP group. In addition, pretreatment mice with NC significantly reduced caspase-3 immunoreactivity induced by CP in testis. Conclusions: This study showed that NC with scavenging free radicals and antiapoptotic properties enable to reduce the side effects of CP in the testicular tissue.

Background Recent studies shows that hyperbaric oxygen (HBO) therapy exerts some protective effects against neural injuries. The purpose of this study was to determine the neuroprotective effects of HBO following sciatic nerve transection (SNT). Methods Rats were randomly divided into five groups ( n  = 14 per group): Sham-operated (SH) group, SH + HBO group, SNT group, and SNT + pre- and SNT + post-HBO groups (100% oxygen at 2.0 atm absolute, 60 min/day for five consecutive days beginning on 1 day before and immediately after nerve transaction, respectively). Spinal cord segments of the sciatic nerve and related dorsal root ganglions (DRGs) were removed 4 weeks after nerve transection for biochemical assessment of malodialdehyde (MDA) levels in spinal cord, biochemical assessment of superoxide dismutase (SOD) and catalse (CAT) activities in spinal cord, immunohistochemistry of caspase-3, cyclooxigenase-2 (COX-2), S100beta (S100ß), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) in spinal cord and DRG. Results The results revealed that MDA levels were significantly decreased in the SNT + pre-HBO group, while SOD and CAT activities were significantly increased in SNT + pre- and SNT + post-HBO treated rats. Attenuated caspase-3 and COX-2 expression, and TUNEL reaction could be significantly detected in the HBO-treated rats after nerve transection. Also, HBO significantly increased S100ß expression. Conclusions Based on these results, we can conclude that pre- and post-HBO therapy had neuroprotective effects against sciatic nerve transection-induced degeneration.

Oxidative stress has a pivotal role in the pathogenesis of diabetic retinopathy (DR). . (JRL) leaf extract has hypoglycemic and antioxidative properties. This study aimed to determine the ameliorative effects of JRL against diabetic retinopathy. The DR rat model was generated by injection of streptozotocin (STZ). A subset of the diabetic rats received JRL or metformin after the onset of hyperglycemia. Histopathology and immunohistochemistry of apoptotic and inflammatory factors were assessed along with biochemical assessments of lipid peroxidation and antioxidant status. Lipid peroxidation level and catalase activity significantly improved after JRL consumption ( <0. 001). Degeneration of the retina attenuated after JRL consumption. Attenuation of the caspase-3, COX-2, PARP, and S100B expression could be detected significantly ( <0. 001) in the JRL-treated rats. While, blood glucose level decreased after JRL consumption ( <0. 001). JRL leaf extract exert protective effects against diabetic retinopathy.

Cisplatin (CP), as an anti-neoplastic drug, causes testicular damage. Zataria multiflora Boiss (ZM), a medicinal plant, has antioxidant and anti-inflammatory properties. The aim of this study was to investigate the effects of ZM against CP-induced testicular toxicity. In this experimental study, thirty-two adult male mice were randomly divided into four groups. The control group received normal saline with oral gavage during 7 days; ZM group received ZM (200 mg/kg) during 7 days by gavage; CP group received CP (10 mg/kg) intraperitoneally (IP) in the 5 day of study; ZM + CP group received ZM during 7 days and CP was injected in 5th day. Sperm parameters, biochemical (MDA, GSH, and PC) levels, serum testosterone levels, and histopathological and immunohistochemical assays of testis were examined one day after the last drug treatment. CP treatment caused significant damage via changed sperm parameters (sperm motility, count, viability rate, and abnormalities), increased oxidative stress (increased MDA and PC levels, and decreased GSH level), histological changes (degeneration, necrosis, arrest of spermatogenesis, congestion, and decrease in thickness of the germinal epithelium, diameter of seminiferous tubules, and Johnsen's Score), decreased serum testosterone level, and increased caspase-3 immunoreactivity. ZM preserved spermatogenesis and mitigated the toxic effects of CP on the testis tissue. In addition, treatment with ZM significantly reduced caspase-3 immunoreactivity. The findings of this study suggest that ZM as a potential antioxidant compound and due to free radicals scavenging activities has a protective effect against CP-induced testicular toxicity.

Deltamethrin (DM) is a synthetic pyrethroid insecticide which can lead to pathological effects in mammals through oxidative stress. On the other hand, virgin olive oil (VOO) is a rich source of phenolic compounds with antioxidants. The aim of the present study was to determine the protective effects of VOO against DM-induced hepatotoxicity. Thirty-six mice were randomly separated into 4 groups: vehicle group, VOO group, DM group, and DM plus VOO group. Immunohistochemistry of PARP, COX-2, and caspase-3 with the biochemical analysis of malondialdehyde and total antioxidant capacity levels were performed in the liver samples 5 weeks after gavaging. Statistical analysis was performed using SPSS, version 15. The data were compared between the groups using the Tukey multiple comparison tests and the analysis of the variance. A P value <0.05 was considered significant. The malondialdehyde level in the liver was increased in the DM group (71.18±0.01), whereas it was significantly (P=0.001) decreased after VOO administration in the DM plus VOO group (39.59±2.43). While the total antioxidant capacity level in the liver was decreased in the DM group (3.05±0.05), it was significantly increased (P=0.03) after VOO administration in the DM plus VOO group (3.95±0.04). A greater expression of caspase-3 (P=0.008), COX-2 (P =0.004), and PARP (P 0.006) could be detected in the DM group, while it was significantly (P=0.009) attenuated in the DM plus VOO group. Also, the degeneration of hepatocytes, which was detected in the DM group, was attenuated after VOO consumption. VOO exerted protective effects against DM-induced hepatotoxicity, which might be associated with its anti-apoptotic, anti-inflammatory, and antioxidative properties.

There is accumulating evidence that Juglans regia L. (GRL) leaf extract has hypoglycemic and antioxidative properties. The present study aimed to investigate the protective effects of GRL leaf extract against diabetic nephropathy (DN). In total, 28 male adult Sprague-Dawley rats were used. The DN rat model was generated by intraperitoneal injection of a single 55 mg/kg dose of streptozotocin (STZ). A subset of the STZ-induced diabetic rats received intragastric administration of GRL leaf extract (200 mg/kg/day) starting 1 week (preventive group) and 4 weeks (curative group) after the onset of hyperglycemia up to the end of the 8th week, whereas other diabetic rats received only isotonic saline (diabetic group) as the same volume of GRL leaf extract. To evaluate the effects of GRL leaf extract on the diabetic nephropathy, various parameters of apoptosis and inflammation were assessed. Statistical analysis was performed using the SPSS software, version 15.0. The data were compared between the groups using the Tukey's multiple comparison test and the analysis of the variance. P values ˂0.05 were considered statistically significant. Fasting blood sugar (FBS) levels (P=0.001) and histopathological changes in the kidney of diabetic rats attenuated after GRL leaf extract consumption. Greater caspase-3 (P=0.004), COX-2 (P=0.008), PARP (P=0.007), and iNOS (P=0.005) expression could be detected in the STZ-diabetic rats, which were significantly (P=0.009) attenuated after GRL leaf extract consumption. In addition, attenuation of lipid peroxidation in the diabetic rats was detected after GRL consumption (P=0.01). GRL leaf extract exerts preventive and curative effects against diabetic nephropathy.