Melatonin and endothelial cell-loaded alginate-fibrin hydrogel promoted angiogenesis in rat cryopreserved/thawed ovaries transplanted to the heterotopic sites.

BACKGROUND: Ischemic niche can promote follicular atresia following the transplantation of cryopreserved/thawed ovaries to the heterotopic sites. Thus, the promotion of blood supply is an effective strategy to inhibit/reduce the ischemic damage to ovarian follicles. Here, the angiogenic potential of alginate (Alg) + fibrin (Fib) hydrogel enriched with melatonin (Mel) and CD144+ endothelial cells (ECs) was assessed on encapsulated cryopreserved/thawed ovaries following transplantation to heterotopic sites in rats. METHODS: Alg + Fib hydrogel was fabricated by combining 2% (w/v) sodium Alg, 1% (w/v) Fib, and 5 IU thrombin at a ratio of 4: 2: 1, respectively. The mixture was solidified using 1% CaCl2. Using FTIR, SEM, swelling rate, and biodegradation assay, the physicochemical properties of Alg + Fib hydrogel were evaluated. The EC viability was examined using an MTT assay. Thirty-six adult female rats (aged between 6 and 8 weeks) with a normal estrus cycle were ovariectomized and enrolled in this study. Cryopreserved/thawed ovaries were encapsulated in Alg + Fib hydrogel containing 100 µM Mel + CD144+ ECs (2 × 104 cells/ml) and transplanted into the subcutaneous region. Ovaries were removed after 14 days and the expression of Ang-1, and Ang-2 was monitored using real-time PCR assay. The number of vWF+ and α-SMA+ vessels was assessed using IHC staining. Using Masson's trichrome staining, fibrotic changes were evaluated. RESULTS: FTIR data indicated successful interaction of Alg with Fib in the presence of ionic cross-linker (1% CaCl2). Data confirmed higher biodegradation and swelling rates in Alg + Fib hydrogel compared to the Alg group (p < 0.05). Increased viability was achieved in encapsulated CD144+ ECs compared to the control group (p < 0.05). IF analysis showed the biodistribution of Dil+ ECs within hydrogel two weeks after transplantation. The ratio of Ang-2/Ang-1 was statistically up-regulated in the rats that received Alg + Fib + Mel hydrogel compared to the control-matched groups (p < 0.05). Based on the data, the addition of Mel and CD144+ ECs to Alg + Fib hydrogel reduced fibrotic changes. Along with these changes, the number of vWF+ and α-SMA+ vessels was increased in the presence of Mel and CD144+ ECs. CONCLUSIONS: Co-administration of Alg + Fib with Mel and CD144+ ECs induced angiogenesis toward encapsulated cryopreserved/thawed ovarian transplants, resulting in reduced fibrotic changes.

Novel Approaches Used in Ovarian Tissue Transplantation for Fertility Preservation: Focus on Tissue Engineering Approaches and Angiogenesis Capacity.

Due to the impact of the modern lifestyle, female infertility has been reduced because of different reasons. For example, in combined chemotherapeutic therapies, a small fraction of cancer survivors has faced different post-complications and side effects such as infertility. Besides, in modern society, delayed age of childbearing has also affected fertility. Nowadays, ovarian tissue cryopreservation and transplantation (OTC/T) is considered one of the appropriate strategies for the restoration of ovarian tissue and bioactivity in patients with the loss of reproductive function. In this regard, several procedures have been considered to improve the efficacy and safety of OTT. Among them, a surgical approach is used to transplant ovaries into the optimal sites, but the existence of ischemic changes and lack of appropriate revascularization can lead to bulk follicular atresia. Besides, the role of OTC/T is limited in women of advanced maternal age undergoing lifesaving chemo-radiation. As a correlate, the development of de novo approaches with efficacious regenerative outcomes is highly welcomed. Tissue engineering shows high therapeutic potentialities to restore fertility in males and females using the combination of biomaterials, cells, and growth factors. Unfortunately, most synthetic and natural materials are at the experimental stage and only the efficacy has been properly evaluated in limited cases. Along with these descriptions, strategies associated with the induction of angiogenesis in transplanted ovaries can diminish the injuries associated with ischemic changes. In this review, the authors tried to summarize recent techniques, especially tissue engineering approaches for improving ovarian function and fertility by focusing on angiogenesis and neovascularization.

Review of ovarian tissue cryopreservation techniques for fertility preservation.

Ovarian failure and ovarian malfunction are among major fertility problems in women of reproductive age (18-35 years). It is known that various diseases, such as ovarian cancer and premature ovarian failure, besides certain treatments, such as radiotherapy and chemotherapy of other organs, can affect the normal process of folliculogenesis and cause infertility. In recent years, various procedures have been proposed for the treatment of infertility. One of the newest methods is the use of cryopreservation ovarian fragments after cancer treatment. According to some studies, this method yields very satisfactory results. Although ovarian tissue cryopreservation (OTC) is an accepted technique of fertility preservation, the relative efficacy of cryopreservation protocols remains controversial. Considering the controversies about these methods and their results, in this study, we aimed to compare different techniques of ovarian cryopreservation and investigate their advantages and disadvantages. Reviewing the published articles may be possible to identify appropriate strategies and improve infertility treatment in these patients.

The effect of fludrocortisone on the uterine receptivity partially mediated by ERK1/2-mTOR pathway.

Implantation of embryos needs endometrial receptivity. Mineralocorticoids is one of the causes influencing the implantation window. This study targeted to evaluation fludrocortisone different properties on endometrial receptivity. The objective of this study was to assess whether treatment with fludrocortisone could impact the expression of diverse genes and proteins that are involved in uterine receptivity in mice. In this study, 40 female adult BALB/c mice were used. The samples were allocated to four groups of ten. Control group (C) received: vehicle; fludrocortisone group (FCA): received 1.5 mg/kg fludrocortisone; PP242 group (PP242): received 30 mg/kg PP242; fludrocortisone+PP242 group (FCA+PP242): received fludrocortisone and PP242. Mice were killed on window implantation day after mating and confirmed pregnancy. The endometrial epithelium of mouse was collected to assess mRNA expression of leukemia inhibitory factor (LIF), mucin-1 (MUC1), heparin-binding epidermal growth factor (HB-EGF), (Msx.1), miRNA Let-7a, and miRNA 223-3p as well as protein expression of extracellular signal-regulated kinase 1/2 (ERK1/2), mammalian target of rapamycin (mTOR), and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) in the uterine using real-time PCR and western blot, respectively. In comparison with the control group, fludrocortisone administration upregulated the expression of LIF, HB-EGF, Msx.1, miRNA Let-7a, ERK1/2, and mTOR in the epithelial endometrium. The PP242-treated group demonstrated a significant rise in the expression of MUC1, miRNA 223-3p and a remarkable decline in ERK1/2 and p-4E-BP1 levels in comparison with the control group. Combination therapy of (FCA+PP242) resulted in a remarkable rise in LIF, Msx-1, HB-EGF, ERK1/2, and mTOR levels, in comparison with the PP242 group. Furthermore, combination therapy of (FCA+PP242) downregulated the expression of MUC1 in comparison with the PP242-treated group. According to the results, fludrocortisone affected uterine receptivity possibly by means of modulating the expression of genes involved in the uterine receptivity and activation of the ERK1/2-mTOR pathway.

Administration of dexamethasone disrupts endometrial receptivity by alteration of expression of miRNA 223, 200a, LIF, Muc1, SGK1, and ENaC via the ERK1/2-mTOR pathway.

Successful implantation of embryos requires endometrial receptivity. Glucocorticoids are one of the factors influencing the implantation window. In this study, 40 female BALB/c mice were used to study the impacts of dexamethasone administration on endometrial receptivity markers during implantation window. The mice mated and were randomly divided into four groups: control (vehicle), dexamethasone (100 µg/kg, IP), PP242 (30 mg/kg, IP), and dexamethasone + PP242 (Dex + PP242). On the Day 4th and 5th of gestation, mice received their respective treatments and were killed on the 5th day. To assess the expression of Muc1, leukemia inflammatory inhibitor (LIF), serum/glucocorticoid-inducible kinase 1 (SGK1), epithelial Na+ channel (ENaC), miRNA 200a, and miRNA 223-3p in the endometrium real-time polymerase chain reaction was performed. Furthermore, using Western blot analysis protein expressions of extracellular signal-regulated kinase 1/2 (ERK1/2), mammalian target of rapamycin (mTOR), and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) were evaluated. Periodic Acid-Schiff staining was used to examine the histomorphological changes of the uterus. According to the results dexamethasone declined the expression of LIF, whereas upregulated expression of Muc1, SGK1, ENaC mRNA, miRNA 200a, and miRNA 223-3p in the endometrium. In addition, PP242, an mTOR inhibitor, induced mRNA expression of Muc1, miRNA200a, and miRNa223-3p whereas it declined the expression of LIF. Moreover, activity of the ERK1/2-mTOR pathway in the endometrial cells was deterred by dexamethasone and PP242. Nonstop epithelium proliferation and elevated surface glycoproteins layer on epithelium of dexamethasone and/or PP242-received groups were divulged through histochemical analysis. According to the above mentioned results, uterine receptivity during implantation period was declined by dexamethasone, at least in part, through modulation of involved genes in endometrial receptivity and inhibition of the ERK1/2-mTOR pathway.

A variation of Musculocutaneous nerve without piercing the coracobrachialis muscle while communicating to the median nerve: A case report and literature review.

INTRODUCTION: Anatomical variations of the peripheral nervous system may have not any clinical signs and symptoms. One of these variations belongs to the Musculocutaneous nerve. However, a good knowledge of nerve pathways and their variations is very important for surgeons in post-traumatic evaluations, exploratory interventions, and/or administration of neuromuscular blocks in axillary region in order to surgical therapies. PRESENTATION OF CASE: This report describes a case of variation of the musculocutaneous nerve which was observed in an old Iranian male cadaver during routine educational dissection (Fig. 1). DISCUSSION AND CONCLUSION: Anatomically, in the axilla region, the Musculocutaneous nerve is originated of the lateral cord of brachial plexus, then, by piercing the coracobrachialis muscle arrives enters to anterior compartment of the arm. But, in the present report, we observed that the Musculocutaneous nerve without piercing the coracobrachialis muscle has arrived in the left arm, then communicated to the Median nerve. To exploratory interventions of the arms for peripheral nerve repair and surgical therapies, a good knowledge of nerve pathways helps to surgeons for preventing possible mistakes during surgery.