Iranian Journal of Basic Medical Sciences
Volume:25 Issue: 9, Sep 2022

Diabetic wounds are the most critical complication in patients with diabetes, which often lead to hospitalization and limb amputations. Diabetic foot ulcers (DFU) is characterized by infections, prolonged inflammation, and a delayed wound healing process. Different types of medical procedures including surgical therapy, drug delivery, stem cell therapy, and wound dressings are used to manage DFU. Bioactive dressings such as hydrogels, nanofiber, and collagens are promising materials that can accelerate the healing process. The wound dressing materials can also be loaded with bioactive molecules like nucleic acids. MicroRNAs (miRNAs) are small non-coding RNA molecules that have recently emerged as regulators of impaired wound healing and could be a target for DFU treatment. miRNA therapeutics can be delivered to the wound region using different delivery systems such as exosomes and nanoparticles. These wound dressings have a high potential for treating diabetic wounds by topical delivery of certain miRNAs in a sustained release manner.

Hypericum perforatum (Hypericaceae), known as Saint John’s wort (SJW), has been used in different systems of traditional medicine such as Chinese traditional medicine, Greek traditional medicine, and Islamic traditional medicine. The plant and its active constituents such as hyperforin and hypericin have a wide range of medicinal uses, particularly as anti-depressant, wound-healing, and antibacterial agents. In recent decades, many clinical trials have been performed to investigate the safety and efficacy of this medicinal plant. However, to the best on our knowledge, there is no comprehensive review article in this regard. In the current study, we aim to have a comprehensive review of the clinical trials of SJW to evaluate its efficacy and safety as well as its application in traditional medicine. Clinical studies investigating the safety, interactions, and efficacy of SJW were identified and summarized, including contributions from 2000 until December 2021. According to the results, these clinical studies were divided into three main categories based on the type of disease: psychiatric, endocrine, and skin problems. Important details of the studies, including the type and duration of the study, the type and percentage of the effective compounds or the extract used, the number of patients, and the obtained results were also discussed. In addition, co-administration and drug interaction of SJW with other drugs were summarized. SJW is a valuable medicinal plant, especially for psychiatric disorders. However, precautions should be taken while administrating the plant.

The Clostridium genus is composed of a large spectrum of heterogeneous bacteria. They are Gram-positive, mostly mesophilic, and anaerobic spore-forming strains. Clostridia are widely distributed in oxygen-free habitats. They are found principally in the soil and intestines of ruminants as normal flora, but also are the cause of several infections in humans. The infections produced by important species in humans include botulism, tetanus, pseudomembranous colitis, antibiotics-associated diarrhea, and gas gangrene. Immunization with toxoid or bacterin-toxoid or genetically modified or other vaccines is a protective way against clostridial infection. Several experimental or commercial vaccines have been developed worldwide. Although conventional vaccines including toxoid vaccines are very important, the new generation of vaccines is an effective alternative to conventional vaccines. Recent advances have made it possible for new vaccines to increase immunogenicity. This review discusses briefly the important species of clostridia in humans, their toxins structure, and vaccine development and usage throughout the world.

Vaccination using inactivated bacteria is one of the most effective ways to protect against EHEC infection. Escherichia coli O157:H7 infections are mainly influenced by Shiga toxins (Stx) and attaching/effacing factors. Among various factors, Stx2B is gaining much attention as a vaccine candidate. Formulating an inactivated bacteria with a suitable adjuvant increases vaccine efficacy and antibody production and can lead to a lasting immune response and protection against O157:H7. 

To assess vaccine efficacy, in this study, we have considered heat and formalin-inactivated bacteria along with chitosan-coated Stx2B/ Stx2B in a mouse model. Ionotropic gelation via tripolyphosphate anions was used to coat Stx2B on chitosan. Subcutaneous injection or oral gavage was used to immunize mice, which were then challenged with E. coli O157:H7. 

Immunity and protection against E. coli  O157:H7 were achieved by all forms of the vaccine. Inactivated E. coli  O157:H7 formulated with chitosan-coated Stx2B effectively evoked humoral and mucosal immune responses. However, minimum shedding appeared with the mice groups orally immunized with formalin-inactivated bacteria sublimated with chitosan-coated Stx2B and heat-inactivated bacteria plus Stx2B in subcutaneous immunization.

Administration of inactivated whole-cell and toxin was synergistic and increased the protection capacity with both parenteral and oral immunization routes.

This study aimed to investigate hypothyroidism’s effects on endometrial receptivity, creating an experimental hypothyroidism model in female rats. 

To induce hypothyroidism in rats of Hipotiroid-ER and Treatment-ER groups, 0.05% 6-propyl-2-thiouracil was freshly added to their drinking water for 8 weeks and then the endometrial receptivity model was applied and sacrificed on the fifth day. In the Treatment-ER group, after sc-administration of 0.8 µg/100 g L-thyroxine for 10 days, the endometrial receptivity model was applied to the rats and sacrificed on the fifth day. 

In the histopathological evaluation, epithelial degeneration, vacuolization, enlargement of the uterine glands, and morphological disorders were observed in the endometrial layer of the Hypothyroid-ER group. However, these pathologies were significantly alleviated in the Treatment-ER group. Integrin β3, integrin αvβ3, LIF, and HOXA10 immune reaction intensities were high in the Control-ER and Treatment-ER groups, while in the Hypothyroid-ER group, integrin β3, integrin αvβ3, LIF, and HOXA10 immunoreactivity intensities were low. Also, while MUC1 immunoreactivity was high in the Hypothyroid-ER group, it was low in the other groups. In biochemical analysis, a significant increase in the TSH and progesterone levels and a significant decrease in the FT4, E2, FSH, and LH levels in the Hypothyroid-ER group compared with the Control-ER group were observed. Also, all hormone levels were significantly ameliorated in the rats of the Treatment-ER group compared with the Hypothyroid-ER group. 

The results obtained showed that hypothyroidism had a significant effect on endometrial receptivity—the histopathological and biochemical changes caused by hypothyroidism in the experimental rat model were ameliorated with L-thyroxine treatment.

Impaired coronary blood flow causes cardiac ischemia. Cellular therapy is a new approach to the treatment of myocardial ischemia. This study aimed to investigate the effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) conditioned with vasopressin on oxidative stress, perivascular collagen, and angiogenesis caused by myocardial infarction (MI) in rats.  We divided 40 male albino Wistar rats into 4 groups; Control group; No intervention; in experimental groups, after it generated induced MI on models, it divided into three groups: Vehicle group (150 μl of cell-free culture medium received); ASC-MI group (6× 106 AD-MSC received) and AVP-ASC-MI group (received 6 × 106 AD-MSC conditioned with 10 nM vasopressin). Then, histologic parameters and anti-oxidant enzymes were evaluated 7 days post-MI cell injection.  Arterial muscle diameter improved and collagen deposition around the coronary arteries decreased in cell-received groups compared with the vehicle group. Malondialdehyde (MDA), catalase (CAT), (GSH) Glutathione, and Total Anti-oxidant Capacity (TAC) parameters were not significantly different between the cells received groups compared with the vehicle group. But the Catalase (CAT) parameter in the ASC-MI group had a significant increase from the control group.  We prepared direct evidence that intramyocardial injection of AD-MSCs reveals the positive cardiac remodeling post-MI in rats, and these useful effects can be more enhanced by administrating injection of conditioned ADSCs with vasopressin.

Ecstasy is a popular recreational psychostimulant with side effects on the central nervous system. This study examined the corpus striatum tissue of adult rats that received ecstasy during the embryonic period for histological and molecular studies.

Rats were divided into control and ecstasy groups. The ecstasy group was given MDMA 15 mg/kg intraperitoneally twice daily at 8-hour intervals on days 7–15 of gestation. At the age of 15 weeks, adult offspring of both groups were examined for learning and memory study by the Morris water maze test. Then, ventral striatum tissue was harvested for TUNEL assay, Nissl staining, and real-time PCR for the expression of the GFAP and CD11b. 

Ecstasy up-regulated the GFAP and CD11b expression in the striatum of offspring (*P˂0.05). Furthermore, the Morris water maze test showed that exposure to ecstasy significantly impaired learning and spatial memory (*P˂0.05). TUNEL assay results did not show any significant change in the number of apoptotic cells in the striatum tissue of ecstasy offspring compared with controls, while Nissl staining showed a significant decrease in the number of neurons in the ecstasy group (*P˂0.05).

Exposure to ecstasy during pregnancy causes long-lasting changes in brain regions underlying learning and memory, including the striatum, and impaired working memory in the offspring. In addition, these data provide the first evidence that exposure to ecstasy during the embryonic period causes a persistent change in the activity of microglial cells and the number of astrocyte cells in the striatum.

Hyperinsulinemia, secondary to insulin resistance, may lead to vascular smooth muscle cell dysfunction. In the present research, we aimed to investigate the effect of Chemokine receptor 8 (CCR8) on angiotensin II (Ang II)-induced dysfunction of vascular smooth muscle cells (VSMCs) and to explore the underlying molecular mechanism. 

The expression of CCR8 was analyzed in diabetics and normal people by RT-PCR and ELISA. CCK-8 assay and transwell were used to explore cell proliferation and migration, and ELISA was used to measure the content of IL-6 and TNF-α. Reactive oxygen species (ROS) kit was employed to measure ROS generation. 

The results revealed that CCR8 was highly expressed in diabetics and Ang Ⅱ-induced VSMCs. Further studies found that interfering with the expression of CCR8 significantly reduced the production of ROS and the levels of inflammatory factors in AngⅡ-induced VSMCs. Interfering with CCR8 increased the glucose uptake induced by AngⅡ+IR. More importantly, inhibition of CCR8 alleviated Ang II-induced dysfunction of VSMCs. Inhibition of CCR8 inactivated the MAPK/NF-κB signaling pathway.

Inhibition of CCR8 attenuates Ang II-induced VSMCs injury by inhibiting the MAPK/NF-κB pathway. CCR8 may be a new biomarker related to hypertension and insulin resistance and is a new target for the treatment of human cardiovascular diseases.

To enhance the efficiency of radiotherapy (RT), implementation of individual-based treatment is essential. In this way, determining individual intrinsic radiosensitivity (IRS) can be useful to achieve minimal adverse effects of RT. The present study aimed to identify IRS of breast cancer (BC) patients through determination of radiation-induced DNA double-strand breaks (DSBs), repair kinetics, and acute normal tissue complications induced by RT.

DSBs induction and its repair kinetics in 50 BC patients’ lymphocytes were analyzed by flow cytometric analysis of H2AX Ser-139 phosphorylation at 30 min, 3 and 24 hr after in vitro irradiation. In vivo skin dosimetry was done by GAFChromic films and acute skin toxicity was scored by radiation oncologists according to the criteria of Radiation Therapy and Oncology Group (RTOG) in all patients with similar prescribed treatment.

The average surface dose for patients ranged from 0.92 to 1.9 Gy and correlation analysis showed no significant relationship with weekly acute skin reactions. Formation of γH2AX after 30 min, slope of dose-response curve and repair kinetics of DSBs after 3 and 24 hr (intrinsic radiosensitivity) were significantly correlated with the RTOG scores following irradiation (clinical radiosensitivity) (r=0.48 and P-value<0.0001, r=0.72 and P-value<0.0001, r=0.48 and P-value<0.001, and finally r=0.53 and P-value<0.001, respectively; (using Pearson’s correlation test). 

Flow cytometric analysis of DNA DSBs by γH2AX measurement has the potential to be developed into a clinical predictor for identifying the overreactor patients prior to RT. Our result suggests that the slope-related quantity based on the linear pattern of the dose-response curve has the merit to predict overreactor patients with a sensitivity of 89% and a specificity of 94%.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), giving rise to the coronavirus disease 2019 (COVID-19), has become a danger to wellbeing worldwide. Thus, finding efficient and safe vaccines for COVID-19 is of great importance. As a basic step amid contamination, SARS-CoV-2 employs the receptor-binding domain (RBD) of the spike protein to lock in with the receptor angiotensin-converting enzyme 2 (ACE2) on host cells. SARS-CoV-2 receptor-binding domain (RBD) is the main human antibody target for developing vaccines and virus inhibitors, as well as neutralizing antibodies. A bacterial procedure was developed for the expression and purification of the SARS-CoV-2 spike protein receptor-binding domain.

In this research study, RBD was expressed by Escherichia coli and purified with Ni-NTA chromatography. Then it was affirmed by the western blot test. The immunogenicity and protective efficacy of RBD recombinant protein were assessed on BALB/c mice. Additionally, RBD recombinant protein was tested by ELISA utilizing sera of COVID-19 healing patients contaminated with SARS-CoV-2 wild type and Delta variation. 

Indirect ELISA was able to detect the protein RBD in serum of the immunized mouse expressed in E. coli. The inactive SARS-CoV2 was detected by antibodies within the serum of immunized mice. Serum antibodies from individuals recovered from Covid19 reacted to the expressed protein.

Our findings showed that RBD is of great importance in vaccine design and it can be used to develop recombinant vaccines through induction of antibodies against RBD.

Non-alcoholic fatty liver disease (NAFLD) is a hepatic manifestation of clinical metabolic syndrome. Insulin resistance is an important factor in the pathogenesis of NAFLD. Ghrelin, widely distributed in peripheral tissues and the central nervous system, plays a vital role in regulating food intake, energy balance, and substance metabolism. In this study, the effect of intracerebroventricular (ICV) injection of ghrelin receptor antagonist on NAFLD was explored.

A rat model of NAFLD was established by feeding a high-fat diet, and a selective ghrelin receptor antagonist [D-Lys-3]-GHRP-6 was injected via ventricular intubation implantation. The serum total cholesterol (TC), triglycerides (TGs), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and hepatic TGs were measured using the colorimetric method. Fasting plasma glucose (FPG) and fasting plasma insulin (FPI) were determined to calculate homeostatic model assessment insulin resistance (HOMA-IR). Hematoxylin-eosin (HE) and Oil Red O staining were conducted to observe the pathological changes and lipid accumulation in the liver. Hosphatidylinositide3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway protein expressions were measured using western blot analysis.

ICV injection of [D-Lys-3]-GHRP-6 significantly reduced serum lipids, transaminase, and HOMA-IR, improved liver injury, and inhibited lipid accumulation in the liver of NAFLD rats. Moreover, ICV injection of [D-Lys-3]-GHRP-6 significantly up-regulated the phosphorylation levels of PI3K/Akt/mTOR signaling protein expressions in the hypothalamus, indicating a significant improvement in hypothalamic insulin resistance.

Blockade of central ghrelin receptor can treat NAFLD possibly via the hypothalamic PI3K/Akt/mTOR signaling pathway to improve insulin resistance.

Exosomes, as nano-sized extracellular vehicles acting as cell-to-cell communicators, are novel promising therapeutics in the area of bone tissue engineering. Moreover, magnetic nanoparticles, whose integration with other appropriate components is viewed as an intriguing approach to strengthen bone tissue engineering efficacy. We investigated the effect of magnetic enriched with exosomes on osteogenic differentiation.

Exosomes were isolated from human adipose-derived mesenchymal stem cells by Exo-spin™ kit (MSC-EX). Alginate (Alg) scaffold containing 1% (w/w) cobalt ferrite nanoparticles (CoFe2O4) was produced. MSC-EX were gently loaded onto Alg and Alg-cobalt ferrite (Alg-CF) scaffolds yielding Alg-EX and Alg-CF-EX scaffolds. The effects of MSC-Ex and magnetic hydrogel composite under an external static magnetic field (SMF) on proliferation and differentiation of MSCs were evaluated by alkaline phosphatase (ALP) activity measurement, alizarin red staining, and energy dispersive X-ray (EDX) analysis.

Our results showed that Alg and Alg-CF scaffolds were not only cytotoxic but also supported AdMSCs proliferation. MSC-EX loading of the scaffolds enhanced AdMSCs proliferation significantly. According to the results, Alg-CF-EX scaffolds under magnetic stimulation exhibited the most potent effect on osteogenic differentiation of cultured AdMSCs as evidenced by higher ALP activity and mineralization.

We provided evidence that the combination of Alg hydrogel, CFNPs, and MSC-EX resulted in the construction of a bone tissue-engineering scaffold that highly supports the osteogenic commitment of MSCs.

The loss of enteric neurons has been shown to be a major cause of slow transit constipation (STC). Gut microbiota and muscularis macrophages (MMs) are associated with the enteric nervous system (ENS) development and gastrointestinal (GI) motility. This study aimed to investigate whether Dioscin (DIO) increased GI motility and inhibited neuron loss by modulating gut microbiota profile, improving inflammation in the ENS microenvironment.

The STC model was established by loperamide. The alteration of the gut microbiota was analyzed by 16S rDNA sequencing. The longitudinal muscle and myenteric plexus (LMMP) from the colon were prepared for flow cytometry, immunofluorescence, western blot, and qRT-PCR. 

DIO increased the stool number, stool water content and shortened whole gut transit time, helped to recover the gut microbial diversity and microbiota community structure, and increased the abundance of Muribaculaceae in STC mice. Compared with the STC group, the number of MMs and the level of the iNOS, IL-6, and TNFα genes were significantly decreased following DIO treatment. Moreover, DIO may increase the number of HuC/D+ neurons per ganglion by up-regulating the BMP2 secreted by MMs and activating the BMP2/p-Smad1/5/9 signaling pathway. Furthermore, the level of excitatory neurotransmitter AchE in colon tissues exhibited a substantial increase in the DIO group. However, the level of inhibitory neurotransmitter VIP was markedly decreased. 

Our results provide that DIO increases GI motility and inhibits neuron loss by modulating gut microbiota profile, improving inflammation in the ENS microenvironment and up-regulating the BMP2 secreted by MMs.

The high resistance rate of Acinetobacter baumannii and the limited number of available antibiotics have prompted a worldwide effort to develop effective antimicrobial agents. Accordingly, identifying single-chain variable fragment antibodies (scFvs), capable of exerting direct antibacterial activity in an immune system-independent manner, may be making immunocompromised patients more susceptible to A. baumannii infections.

To isolate bactericidal scFvs targeting A. baumannii, we panned a large human scFv phage display library against whole-cell extensively drug-resistant (XDR) A. baumannii strains grown as biofilm or cultured with human blood or human peripheral blood mononuclear cells plus plasma. The binding of scFv-phages to A. baumannii was assessed by the dot-blot assay. Soluble scFvs, derived from the selected phages, were assessed based on their ability to bind and inhibit the growth of A. baumannii. 

Five phage clones showed the highest reactivity toward A. baumannii. Among five soluble scFvs, derived from positive phage clones, two scFvs, EB211 and EB279, had high expression yields and displayed strong binding to A. baumannii compared with the controls. Moreover, XDR A. baumannii strains treated with positively-charged scFvs, including EB211, EB279, or a cocktail of EB211 and EB279 (200 µg/ml), displayed lower viability (approximately 50%, 78%, and 40% viability, respectively) compared with PBS-treated bacteria.

These results suggest that combining last-resort antibiotics with bactericidal scFvs could provide promising outcomes in immunocompromised individuals with A. baumannii infections.

Therapeutic strategies that facilitate extinction are promising in the treatment of post-traumatic stress disorder (PTSD). Brain-derived neurotrophic factor (BDNF) has a crucial role in neural plasticity, a process needed for the retention of fear extinction. In this study, we investigated the effects of local administration of a histone deacetylase (HDAC) inhibitor, sodium butyrate (NaBu), on BDNF transcription and behavioral markers of extinction in the single prolonged stress (SPS) model of PTSD. 

NaBu was infused into the infralimbic (IL) subregion of the medial prefrontal cortex (mPFC) of male rats. The freezing response was recorded as the criterion to assess fear strength on the day of extinction as well as 24 hr later in the retention test. Other behavioral tests were also measured to evaluate the anxiety level, locomotor activity, and working memory on the retention day. HDAC activity and BDNF mRNA expression were evaluated after the behavioral experiments. 

NaBu facilitated the recall of fear extinction in SPS rats (P<0.0001). SPS rats had higher HDAC activity (P<0.0001) and lower BDNF expression (P<0.05) than non-SPS animals. Also, anxiety was higher in the SPS group (P<0.0001), but locomotor activity (P=0.61) and working memory (P=0.36) were not different between SPS and Non-SPS groups.

Our findings provide evidence that the mechanism of action of NaBu in the improvement of extinction recall is mediated, in part, by enhancing histone acetylation and reviving BDNF expression in IL.

Type 1 diabetes mellitus is a common autoimmune and multifactorial disorder. Researchers have been interested in making a favorable islet-like tissue model for the treatment of diabetes. The main objective of this study was to determine the effects of the spleen extracellular matrix (S-ECM) on the function of the MIN6 cell line (a β-cell model). 

In this experimental research, Wistar rat spleens were decellularized by sodium dodecyl sulfate (SDS) and Triton X-100. S-ECM was characterized by histological assessments, scanning electron microscopy, determination of residua DNA, and examination of the mechanical tensile property. Then, MIN6 cells were seeded on S-ECM scaffold. Glucose-stimulated insulin secretion and mRNA expression of insulin-related genes were examined to confirm the function of the cells. 

The main components of S-ECM such as collagen and glycosaminoglycan remained after decellularization. Furthermore, very low residual DNA and appropriate mechanical behavior of S-ECM provided an ideal extracellular microenvironment for the MIN6 cells. GSIS results showed that the seeded cells in S-ECM secreted more insulin than the traditional two-dimensional (2D) culture. The expression of specific insulin-related genes such as PDX-1, insulin, Maf-A, and Glut-2 in the recellularized scaffold was more significant than in the 2D traditional cultured cells. Also, MTT assay results showed that S-ECM were no cytotoxic effects on the MIN6 cells. 

These results collectively have evidenced that S-ECM is a suitable scaffold for stabilizing artificial pancreatic islands.