Trehalose and carnosic acid induced LC3I, LC3II ratio, P62 down-regulation and cleaved caspase 3 expression in neural stem cells.

BACKGROUND: Using neural stem cells (NSCs) in cell therapy and regenerative medicine is a growing knowledge. In this study, the protective role of carnosic acid and trehalose against H2O2-induced oxidative stress in autophagy induction and apoptosis inhibition in NSCs was investigated. MATERIAL AND METHODS: The bone marrow stromal cells (BMSCs) were isolated from the femur of the rat and differentiated into NSCs using basic fibroblast and epidermal growth factors (bFGF and EGF), and B27 serum free media. To evaluate the autophagy, the P62 protein was assessed by immunocytochemistry and LC3II / LC3I ratio by Western blotting. Further, we used 3-Methyladenine (3-MA), a widely used autophagy inhibitor to study whether combined treatment of 3-MA with carnosic acid and trehalose modulates autophagy in NSCs. For studying apoptosis, the cleaved caspase-3 protein was evaluated. Carnosic acid and trehalose increased the survival of the NSCs. RESULTS: The H2O2 decreased the autophagy and induced apoptosis with increasing time during 24 hours, however, a pre-treatment with 2 µM carnosic acid and trehalose 3 % induced the autophagy proteins (while increasing the LC3II / LC3I ratio and decreasing the P62) and decreased the apoptosis (while decreasing the expression of the cleaved caspase-3). The results showed that the carnosic acid and trehalose increased the survival of NSCs against the oxidative stress caused by H2O2, decreased apoptosis, and induced autophagy. CONCLUSION: Due to the carnosic acid and trehalose unique properties and its low toxicity, it can be used as an agent in cellular transplantation for reducing oxidative stress and inducing autophagy (Fig. 4, Ref. 37).

Cerebrospinal fluid and blood exosomes as biomarkers for amyotrophic lateral sclerosis; a systematic review.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a progressive and fatal motor neuron disease. Due to the limited knowledge about potential biomarkers that help in early diagnosis and monitoring disease progression, today's diagnoses are based on ruling out other diseases, neurography, and electromyography examination, which takes a time-consuming procedure. METHODS: PubMed, ScienceDirect, and Web of Science were explored to extract articles published from January 2015 to June 2023. In the searching strategy following keywords were included; amyotrophic lateral sclerosis, biomarkers, cerebrospinal fluid, serum, and plama. RESULTS: A total number of 6 studies describing fluid-based exosomal biomarkers were included in this study. Aggregated proteins including SOD1, TDP-43, pTDP-43, and FUS could be detected in the microvesicles (MVs). Moreover, TDP-43 and NFL extracted from plasma exosomes could be used as prognostic biomarkers. Also, downregulated miR-27a-3p detected through exoEasy Maxi and exoQuick Kit in the plasma could be measured as a diagnostic biomarker. Eventually, the upregulated level of CORO1A could be used to monitor disease progression. CONCLUSION: Based on the results, each biomarker alone is insufficient to evaluate ALS. CNS-derived exosomes contain multiple ALS-related biomarkers (SOD1, TDP-43, pTDP-43, FUS, and miRNAs) that are detectable in cerebrospinal fluid and blood is a proper alternation. Exosome detecting kits listed as exoEasy, ExoQuick, Exo-spin, ME kit, ExoQuick Plus, and Exo-Flow, are helpful to reach this purpose.

The Effect of bisphenol A and Photobiomodulation Therapy on Autophagy-Related Genes Induction in Adipose Tissue-Derived Stem Cells.

Introduction: As adipose tissue-derived stem cells (ADSCs) can divide rapidly and be prepared non-invasively, they have extensively been used in regenerative medicine. On the other hand, a new method of therapy, known as photobiomodulation (PHT), has been used to treat many diseases, such as inflammatory conditions, wound healing and pain. Besides, exposure to chemical substances such as bisphenol A (BPA), at low levels, can lead to autophagy. This study investigated the effects of BPA and PHT on the expression of autophagy-related genes, including LC3, NRF2, P62, in rat ADSCs as a model. Methods: ADSCs isolation and purification were confirmed by immunocytochemistry (ICC). The cells were then treated with different concentrations of BPA and also subjected to PHT. Reverse transcription polymerase chain reaction (RT-PCR) was used for the evaluation of LC3, NRF2 and P62 gene expressions. Oil red O staining was used for adipogenic vacuole formation. Result: ICC showed that the isolated cells were CD 49-positive but CD 31 and CD 34-negative. The viability test indicated that the number of live cells after 24 hours in the BPA groups at concentrations of 0, 1, 50, 100 and 200 µM was 100%, 93%, 81%, 72%, and 43% respectively. The difference in cell viability between groups 50, 100 and 200 µM was significant as compared with the control groups (P < 0.05). Moreover, in the group with 1 µM concentration of BPA, the expressions of LC3, NRF2 and P62 genes were upregulated. However, in the treatment group at the concentration of 200 µM of BPA, the LC3 gene was expressed, but NRF2 and P62 genes were downregulated. Conclusion: BPA and PHT induce autophagy and adiposeness in ADSCs in a dose-dependent manner.

Isolation and Culture of Embryonic Mouse Neural Stem Cells.

Neural stem cells (NSCs) are multipotent and can give rise to the three major cell types of the central nervous system (CNS). In vitro culture and expansion of NSCs provide a suitable source of cells for neuroscientists to study the function of neurons and glial cells along with their interactions. There are several reported techniques for the isolation of neural stem cells from adult or embryo mammalian brains. During the microsurgical operation to isolate NSCs from different regions of the embryonic CNS, it is very important to reduce the damage to the brain cells to obtain the highest ratio of live and expandable stem cells. A possible technique for stress reduction during isolation of these cells from the mouse embryo brain is the reduction of surgical time. Here, we demonstrate a developed technique for rapid isolation of these cells from the E13 mouse embryo ganglionic eminence. Surgical procedures include harvesting E13 mouse embryos from the uterus, cutting the frontal fontanelle of the embryo with a bent needle tip, extracting the brain from the skull, microdissection of the isolated brain to harvest the ganglionic eminence, dissociation of the harvested tissue in NSC medium to gain a single cell suspension, and finally plating cells in suspension culture to generate neurospheres.

Incidence of various cardiac arrhythmias and conduction disturbances due to high dose intravenous methylprednisolone in patients with multiple sclerosis.

BACKGROUND: High-dose intravenous methylprednisolone is the most common therapeutic modality to treat acute exacerbations in multiple sclerosis (MS). Various cardiac arrhythmias have been reported during corticosteroid pulse therapy. This study was conducted to detect cardiac rhythm changes in patients with MS while receiving high dose methylprednisolone. METHODS: We enrolled 52 consecutive MS patients with acute relapse to perform cardiac monitoring 4h before, during and 18 h after infusion of 1000 mg intravenous (IV) methylprednisolone. RESULTS: Sinus tachycardia was the most common change in cardiac rhythms before, during, and after corticosteroid pulse therapy. Up to 41.9% of the patients, developed sinus bradycardia after pulse infusion. Sinus arrest and sinus exit block were observed in 12 patients. Atrial fibrillation and ventricular tachycardia were observed in three patients and one patient, respectively. The most important cardiac arrhythmias including ventricular tachycardia, sinus arrest, and sinus exit block, were correlated with smoking and more commonly observed during 12h post infusion. Sinus bradycardia and atrial fibrillation were detected more commonly in patients with history of urinary dysfunction. CONCLUSION: High dose intravenous prednisolone might cause different types of arrhythmias in MS patients. Cigarette smokers and patients with autonomic disturbances like sphincter and bowel problems have more chance to develop arrhythmias while receiving high dose steroids.