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Tacrolimus, a widely used immunosuppressive drug for preventing graft rejection following organ transplantation, was reported to develop neurotoxic side effects ranging from mild to severe symptoms in the literature. Rats were randomly divided into three groups as control and 2-week and 3-week treatment groups and received a 2 mg/kg/day tacrolimus by oral gavage. Animals were sacrificed and sciatic nerves obtained from all groups were fixed and processed for light and electron microscopic investigations. The myelinated fiber diameter, axon diameter, G-ratio (axon diameter/myelinated fiber diameter), and myelin thickness were also determined. The data obtained in the control and tacrolimus-treated groups were compared.The control group sciatic nerve fascicles showed normal morphology with myelinated and unmyelinated fibers. Experimental groups exhibited axonal dilatation, irregularly thickened and vacuolated myelin sheaths with separation of myelin layers. The morphometric analysis showed that the myelinated fibers of the 2-week tacrolimus-treated group displayed a moderate increase in the myelin thickness and axon and fiber diameter in comparison with the control and 3-week tacrolimus-treated groups. The G-ratio was found to be in normal range in all groups and there were no statistically significant difference.The present study indicates that the treatment with tacrolimus may produce a mild degenerative change but prolonged drug administration for 3 weeks led to improvement in morphometric and morphologic data and the normal G-ratio values, suggesting that the regeneration capacity of the myelinated fibers maintains their normal function to transmit nerve impulses.

Background and Objectives: The HUC-HEART Trial (ClinicalTrials.gov Identifier: NCT02323477) was a controlled, prospective, phase I/II, multicenter, single-blind, three-arm randomized study of intramyocardial delivery of human umbilical cord-derived mesenchymal stromal cells (HUC-MSCs) combined with coronary artery bypass-grafting (CABG) in patients with chronic ischemic cardiomyopathy (CIC). The trial aimed to assess (i) the safety and the efficacy of cell transplantation during one-year follow-up, (ii) to compare the efficacy of HUC-MSCs with autologous bone-marrow- derived mononuclear cells (BM-MNCs) in the same clinical settings., Methods and Results: Fifty-four patients who were randomized to receive HUC-MSCs (23×10 6 ) (n=26) or BM-MNCs (70×10 7 ) (n=12) in combination with CABG surgery. The control patients (n=16) received no cells/vehicles but CABG intervention. All patients were screened at baseline and 1, 3, 6, 12 months after transplantation. Forty-six (85%) patients completed 12 months follow-up. No short/mid-term adverse events were encountered. Decline in NT-proBNP (baseline∼ 6 months) in both cell-treated groups; an increase in left ventricular ejection fraction (LVEF) (5.4%) and stroke volume (19.7%) were noted (baseline∼6 or 12 months) only in the HUC-MSC group. Decreases were also detected in necrotic myocardium as 2.3% in the control, 4.5% in BM-MNC, and 7.7% in the HUC-MSC groups. The 6-min walking test revealed an increase in the control (14.4%) and HUC-MSC (23.1%) groups., Conclusions: Significant findings directly related to the intramyocardial delivery of HUC-MSCs justified their efficacy in CIC. Stricter patient selection criteria with precisely aligned cell dose and delivery intervals, rigorous follow-up by detailed diagnostic approaches would further help to clarify the responsiveness to the therapy.

Purpose: Chemical fixation is a critical step to retaining cellular targets as naturally as possible. Recent developments in microscopy allow sophisticated detection and measuring techniques with which spatio-temporal molecular alterations are conceivable. In this study, we compare two members of aldehyde fixatives [i.e., glyoxal (Gly) and paraformaldehyde (PFA)] to determine whether Gly, a less toxic dialdehyde fixative that is considered to retain immunoreactivity could provide a successful and consistent cell fixation in favor of PFA in various cell preparations and types., Methods: We document the fixation competence of Gly and PFA side-by-side (with or without Triton X-100 permeabilization) in live- and fixed-cell preparations in mouse oocytes, embryos, and human somatic cells (human umbilical cord-derived mesenchymal stromal cells) using protein quantification by Western blot assay and super-resolution microscopy., Results: Although Gly seemed to act faster than PFA, catastrophic consequences were found not acceptable, especially in oocytes and embryos. Due to cell lysate and immunocytochemistry surveys, it was obvious that PFA is superior to Gly in retaining cellular proteins in situ with little/no background staining. In many samples, PFA revealed more reliable and consistent results regarding the protein quantity and cellular localization corresponding to previously defined patterns in the literature., Conclusion: Although the use of Gly is beneficial as indicated by previous reports, we concluded that it does not meet the requirement for proper fixation, at least for the tested cell types and proteins. However, PFA alone with no addition of TX displayed a significant cytoplasmic loss by generating membrane blebs during fixation.

Aim: To demonstrate progression of acute and chronic endocrinopathies in a kaolin-induced hydrocephalus model using light microscopy., Material and Methods: Adult male Sprague-Dawley rats (n = 48) were divided into six groups. Hydrocephalus was induced by intracisternal injection of kaolin solution in the acute and chronic kaolin groups, whereas an identical volume of sterile saline was injected into the sham groups., Results: Somatotropic cell concentrations were lower in the kaolin groups compared with their controls, but there was no difference in somatotropic cell concentration between the acute and chronic kaolin groups. Corticotropic cell concentrations were higher in the acute kaolin and sham groups compared with acute controls. Thyrotropic cell numbers were higher in the acute sham and kaolin groups compared with their controls, and although thyrotropic cell concentations were higher in the acute kaolin group than the acute sham group. No differences were observed between the acute and chronic controls and sham and kaolin groups regarding mammotropicand gonadototropic cell concentations., Conclusion: Somatotropic cells are most affected by hydrocephalus that causes pituitary dysfunction, and this effect was more prominent under acute and chronic phases.

Background: The HUC-HEART Trial is a clinical study of intramyocardial delivery of current Good Manufacturing Practice (cGMP)-grade human umbilical cord multipotent stromal cells (HUC-MSCs) in ischemic cardiomyopathy where 2 × 10 7  cells are administered to peri-infarcted myocardium. Prior to the onset of the trial, we aimed to optimize the transport/storage conditions for obtaining the highest cell viability and proliferation rate of cells to be transplanted., Methods: Cells were tested after being transported in phosphate-buffered saline (PBS) or Ringer's lactate-based (RL) transport media supplemented with human serum albumin (HSA) and/or hydroxyethyl starch (HES) at two temperatures (2-10°C or 22-24°C)., Results: The effects of transport conditions on cell viability following 6 h were found highest (93.4 ± 1.5) in RL-based media at 2-10°C. Karyotypes were found normal upon transportation in any of the formulations and temperatures. However, the highest proliferation rate was noted (3.1-fold increase) in RL (1% HSA) media at 2-10°C over 6 days in culture. From that point, RL (1% HSA) media at 2-10°C was used for further experiments. The maximum cell storage time was detected around 24 h at 2-10°C. Extended storage periods resulted in a decrease in cell viability but not in MSC marker expression. An increase in actin quantity was detected in hypoxia (5% O 2 ) groups in early culture days; no difference was noted between hypoxic versus normoxic (21% O 2 ) conditions in later days., Discussion: The overall results suggest that non-commercial, simple media formulations with extended storage intervals at 2-10°C temperatures are capable of retaining the characteristics of clinical-grade HUC-MSCs. The above findings led us to use RL (1% HSA) media at 2-10°C for transport and storage in the HUC-HEART Trial; 23 patients received HUC-MSCs by August 2018; no adverse effects were noted related to cell processing and transplantation., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Purpose: Even with 86 live births reported globally so far, the mechanism of primordial follicle loss following autotransplantation of the frozen-thawed ovarian tissue needs further evaluation. Pten, Tsc1, p27, and Amh are the inhibitor proteins that play crucial roles in suppressing the transition from the primordial follicle to primary state, maintaining the primordial follicle reserve. In this study, we aimed to evaluate whether the expression patterns of these proteins change and it may be related to the global primordial follicle loss after autotransplantation of the frozen-thawed ovarian tissue., Methods: Four groups were established in rats: fresh-control, frozen/thawed, fresh-transplanted, and frozen/thawed and transplanted. After slow freezing and thawing process, two ovarian pieces were transplanted into the back muscle of the same rat. After 2 weeks, grafts were harvested, fixed, and embedded into the paraffin block. Normal and atretic primordial/growing follicle count was performed in all groups. Ovarian tissues were evaluated for the dynamic expressions of the Pten, Tsc1, p27, and Amh proteins using immunohistochemistry, and H-score analyses were done., Results: Ovarian tissue cryopreservation does not change the expression patterns of inhibitory proteins that control ovarian reserve. Both in fresh and frozen/thawed autotransplanted groups, the expression of inhibitory proteins and Amh decreased significantly in primordial follicles and in growing follicles, respectively. In control group and in frozen/thawed group, primordial follicle counts were similar but decreased by almost half in both fresh-transplanted and frozen/thawed and transplanted groups., Conclusions: One of the causes of primordial follicle loss after transplantation of ovarian graft may be decreased expression of the inhibitory proteins that guard the ovarian reserve and transplantation itself seems to be the major cause for disruption of inhibitory molecular signaling. Our findings highlight important molecular aspects for future clinical applications for fertility preservation in humans.

The advances and success of umbilical cord-derived mesenchymal stromal cells (UC-MSCs) in experimental disease animal models have fueled the development of targeted therapies in humans. The therapeutic potential of allogeneic transplantation of UC-MSCs has been under examination since 2009. The purpose of this systematic analysis was to review the published results, limitations and obstacles for UC-MSC transplantation. An extensive search strategy was applied to the published literature, 93 peer-reviewed full-text articles and abstracts were found published by early August 2017 that investigated the safety, efficacy and feasibility of UC-MSCs in 2001 patients with 53 distinct pathologies including many systemic/local, acute/chronic conditions. Few data were extracted from the abstracts and/or Chinese-written articles (n = 7, 8%). Importantly, no long-term adverse effects, tumor formation or cell rejection were reported. All studies noted certain degrees of therapeutic benefit as evidenced by clinical symptoms and/or laboratory findings. Thirty-seven percent (n = 34) of studies were found published as a single case (n = 10; 11%) or 2-10 case reports (n = 24; 26%) with no control group. Due to the nature of many stem cell-based studies, the majority of patients also received conventional therapy regimens, which obscured the pure efficacy of the cells transplanted. Randomized, blind, phase 1/2 trials with control groups (placebo-controlled) showed more plausible results. Given that most UC-MSC trials are early phase, the internationally recognized cell isolation and preparation standards should be extended to future phase 2/3 trials to reach more convincing conclusions regarding the safety and efficacy of UC-MSC therapies., (Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.)