Efectos funcionales e histológicos posteriores al implante de células madre pluripotentes en un modelo murino con esfinterotomía

Introduction and aimsFecal incontinence is a disabling condition with devastating consequences for the patients. Medical and surgical options are not very satisfactory, reason by which regenerative medicine has been considered in this field. In the present research, we analyzed functional and histologic effects after implanting pluripotent stem cells (PSCs) in a murine model with sphincterotomy.Materials and methodsFemale Wistar rats were subjected to sphincterotomy and divided into three groups. Group 1 (control group) was treated with 300 μL of balanced saline solution and group 2 (late treatment) and group 3 (early treatment) received 50,000 PSCs resuspended in 300 μL of balanced saline solution. All animals were evaluated through high-resolution anorectal manometry 24 hours before and after sphincterotomy and every month for three months. Finally, the rats were euthanized and histopathologic sections from the anal canal were obtained.ResultsAll groups showed a decrease in resting anal pressure and squeeze anal pressure 24 hours after sphincterotomy. At the third month, higher anal pressures in the groups treated with PSCs were detected. Regarding the histologic effects, the microscopic architecture was restored and there was a significant decrease in the inflammatory response in the groups treated with PSCs.ConclusionPSCs implantation improves anal tone, as well as histologic structure, presenting better regenerative results when implanted as early treatment.     

Regenerative medicine of pancreatic islets

The pancreas became one of the first objects of regenerative medicine,since other possibilities of dealing with the pancreatic endocrine insufficiency were clearly exhausted.The number of people living with diabetes mellitus is currently approaching half a billion,hence the crucial relevance of new methods to stimulate regeneration of the insulin-secreting β-cells of the islets of Langerhans.Natural restrictions on the islet regeneration are very tight;nevertheless,the islets are capable of physiological regeneration via β-cell self-replication,direct differentiation of multipotent progenitor cells and spontaneous α-to or δ-to β-cell conversion(trans-differentiation).The existing preclinical models of β-cell dysfunction or ablation(induced surgically,chemically or genetically) have significantly expanded our understanding of reparative regeneration of the islets and possible ways of its stimulation The ultimate goal,sufficient level of functional activity of β-cells or their substitutes can be achieved by two prospective broad strategies β-cell replacement and β-cell regeneration.The "regeneration" strategy aims to maintain a preserved population of β-cells through in situ exposure to biologically active substances that improve β-cell survival,replication and insulin secretion,or to evoke the intrinsic adaptive mechanisms triggering the spontaneous non-β-to β-cell conversion.The "replacement" strategy implies transplantation of β-cells(as non-disintegrated pancreatic material or isolated donor islets) or β-like cells obtained ex vivo from progenitors or mature somatic cells(for example,hepatocytes or a-cells) under the action of small-molecule inducers or by genetic modification.We believe that the huge volume of experimental and clinical studies will finally allow a safe and effective solution to a seemingly simple goal-restoration of the functionally activeβ-cells, the innermost hope of millions of people globally.     

Ex Vivo Radiation Leads to Opposing Neurite Growth in Whole Ganglia vs Dissociated Cultured Pelvic Neurons

BackgroundProstatic radiation therapy (RT) often causes erectile dysfunction (ED) and the mechanisms governing RT-induced ED are unclear with a lack of therapeutic strategies.AimTo determine the effects of ex vivo RT on major pelvic ganglion (MPG) neuron survival, and neurite growth in whole vs dissociated culture.MethodsMPGs were removed and irradiated (0 or 8 Gy) from male Sprague Dawley rats. For dissociated culture, MPG neurons were digested in collagenase/dispase and cultured on coverslips. Immunofluorescent staining for beta-tubulin III (TUBB3; neuron marker), neuronal nitric oxide synthase (nNOS; nitrergic marker), tyrosine hydroxylase (TH; sympathetic marker), and terminal deoxynucleotidyl transferase dUTP nick end labeling assessed neurite length, branching, autonomic neuron density, and apoptosis. For whole organ culture, MPGs were grown in Matrigel. Gene expression of apoptotic markers (caspase 1, 3), TUBB3, nNOS, TH, and Schwann cells (Sox10, Krox20, glial fibrillary acid protein) was measured in whole organ cultured MPGs by quantitative polymerase chain reaction.OutcomesAfter 72 hours, neurite length, branching, autonomic neuron density, and apoptosis were assessed, and gene expression was measured.ResultsRT increased apoptosis in dissociated neurons measured by terminal deoxynucleotidyl transferase dUTP nick end labeling (P < .001) and whole MPG culture via upregulation of caspase 3 gene expression (P < .05). Nitrergic neurons were markedly decreased in irradiated dissociated culture (P < .05), while nNOS gene expression was upregulated in irradiated whole organ culture (P < .05). The proportion of dissociated sympathetic neurons and whole organ TH gene expression remained unchanged after RT. Interestingly, RT dissociated neurites were 22% shorter than controls, while RT whole organ neurites were 15% longer than controls (P < .01). MPG Schwann cells markers (Sox10, Krox20) were elevated after RT in whole organ culture.Clinical TranslationProstatic RT leads to increased neuronal cell death and less erectogenic nitrergic neurons contributing to ED.Strengths & LimitationsThe advantages of dissociated neuron culture include distinct neurites which are easily measured for apoptosis, length/branching, and specific neuron types. In contrast, whole MPG culture is advantageous as it contains all the supporting cells present in vivo.ConclusionThe 2 different culture methods demonstrated opposing neurite growth after RT indicating the importance of supporting cell network to promote pelvic neuron neuritogenesis and survival following RT.Randolph JT, Pak ES, Koontz BF, et al. Ex Vivo Radiation Leads to Opposing Neurite Growth in Whole Ganglia vs Dissociated Cultured Pelvic Neurons. J Sex Med 2020;17:1423–1433.     

Wnt10b 诱导再生毛囊的表达特性研究

目的：研究 Wnt10b 诱导再生毛囊的表达特性及诱导作用机制。方法 HEK-293细胞内扩增并用氯化铯梯度离心纯化 Wnt10b 过表达腺病毒及对照腺病毒，皮内注射至 C57BL/6J 小鼠背部皮肤，在处理后2.5、5、7、9、14、28 d 时取材，HE 染色及免疫组化染色观察毛囊结构特征、信号通路表达特征及增殖特性。结果HE 染色发现，AdWnt10b 处理组从第5天开始出现新生毛囊结构，正常生长，第28天左右进入退化期。免疫组化染色发现，AdWnt10b 处理组从处理后5 d 开始新生毛囊具有 AE15表达，随着毛囊生长而增加，至处理后28 d开始减少。在 AdWnt10b 处理后5 d，观察到β连环素的核表达，Lef1特异性表达于毛芽和毛母质部位，且全为核表达。在 AdWnt10b 处理后28 d，Lef1表达减弱。AdWnt10b 处理后2.5 d 即可见 Ki67表达于表皮和毛囊外根鞘。处理后2.5、7、9、14 d 均在隆突区见到 Ki67的表达；从处理后7 d 开始，Ki67表达于毛母质细胞。结论Wnt10b 诱导的再生毛囊具有正常的毛囊结构，Wnt10b 激活了经典 Wnt 信号通路，其作用的靶细胞是毛囊干细胞及其子代细胞。     

胆汁酸在梗阻性黄疸小鼠肝部分切除术后肝再生中作用的研究

目的：探讨胆汁酸（CA）在梗阻性黄疸（OJ）小鼠肝部分切除术后肝再生中的作用及机制。 方法：180只健康雄性小鼠随机均分为6组,分别行假手术（对照组）、胆总管结扎（OJ组）、胆总管结扎并于7 d加行外引流（ED组）、胆总管结扎+0.2%CA灌胃并7 d后加行外引流（ED+0.2%CA组）、胆总管结扎+1%CA灌胃并于7 d后加行外引流（ED+1%CA组）、胆总管结扎并于7 d后加行内引流（ID组）,各组分别于实验第14天行70%肝切除,且各外引流组改行内引流。检测各组肝切除术后不同时间点肝再生率与肝组织增殖细胞核抗原Ki-67表达、叉状头盒M1b基因（Foxm1b）mRNA相对表达、成纤维细胞生长因子受体4（FGFR4）蛋白表达,并观察部分组肝细胞原位凋亡情况。 结果：除对照组外,肝再生率、肝组织Ki-67阳性表达率、Foxm1b mRNA及FGFR4蛋白表达在其余各组均由高到低依次为：ID组>ED+0.2%CA组>ED组>OJ组>ED+1%CA组,组间差异均有统计学意义（均P<0.05）;ID组与对照组间各指标差异均无统计学意义（均P>0.05）;肝细胞凋亡率由高到低依次为：ED+1%C组>ED组>ED+0.2%CA组>对照组,组间差异均有统计学意义（均P<0.05）。 结论：内引流通过减少内源性CA的丢失有利于肝切除后肝再生;外源性低浓度CA可以恢复外引流引起的肝再生障碍,可能与其上调Foxm1b与FGFR4的表达从而促进肝再生有关。     

Evaluating the effect of polytetrafluoroethylene and extractum cepae-heparin-allantoin gel in peripheral nerve injuries in a rat model

BACKGROUND:Peripheral nerves can be injured by congenital, mechanical, thermal or chemical causes. Peripheral nerve injuries are increasing in frequency, particularly in countries that are becoming more industrialized. Nerve and extremity injuries result in work loss and high treatment costs, and can lead to separation of patients from their social environment. Failure of nerve repair causes muscle functional losses, sensory losses and painful neuropathies.OBJECTIVES:To compare the effects of condensed polytetrafluoroethylene (cPTFE) and cPTFE-extractum cepae-heparin-allantoin (cPTFE-EHA) gel compound on nerve and functional recovery, and the prevention of adhesion and scar tissue formation after total peripheral nerve injury repaired by primary suture in a rat model.RESULTS:cPTFE alone and cPTFE-EHA gel was found to provide better functional recovery and nerve regeneration compared with primary repair only. In the macroscopic evaluation, the cPTFE-EHA gel was found to have no negative effect on wound healing and, despite increasing extra-neural scar tissue and adhesions, it had no negative effect on nerve function; in addition, it facilitated functional recovery.CONCLUSIONS:Compared with the cPTFE application alone, the application of perineural cPTFE-EHA gel during peripheral nerve surgery appeared to provide better functional recovery without causing any significant changes in epineural and extraneural scar tissue formation.     

The eternal tooth germ is formed at the apical end of continuously growing teeth

Rodent incisors are known to be continuously growing teeth that are maintained by both the cell-proliferation at the apical end and the attrition of the incisal edge. This type of tooth had a special epithelial structure for the maintenance of stem cells, showing the bulbous epithelial protrusion at the apical end. The morphological transition of the epithelial-mesenchymal compartment by serial transverse sections of the apical end toward the incisal direction is likely to reflect the development of the tooth germ in the prenatal stage. Based on the present histological and previous molecular biological studies, the special structure at the apical end is obviously different from the cervical loop giving rise to Hertwig's epithelial root sheath (HERS), in human, mouse and rat molar tooth germs. Hence, we propose a new concept that the eternal tooth bud producing various dental progeny is formed at the apical end of continuously growing teeth, and a new term "apical bud" for indicating this specialized epithelial structure. Furthermore, BrdU labelling analysis suggested that the guinea-pig molars, which were continuously growing teeth, also possessed plural specific proliferative regions and "apical bud" at the apical end.     

膜引导组织再生术与自体静脉桥接法修复面神经缺损的实验研究

目的观察并比较膜引导组织再生术和静脉桥接法修复面神经的作用和效果。方法分别采用膜引导组织再生术和自体静脉桥接法对家兔进行面神经缺损修复,以自体神经移植为对照,通过电生理学和组织学方法,观察和比较两种修复方法的效果。结果两种方法都具有修复面神经缺损的作用,但效果存在一定差别。膜引导组织再生组,术后3个月时两侧面神经干传导速度之差值为(2.10±1.2)m/s,而静脉桥接组为(6.80±1.4)m/s,神经移植组为(2.16±1.6)m/s。组织学上,膜引导组织再生组,术后3个月神经纤维排列整齐,延续连贯;静脉桥接组,神经纤维排列不整齐,结缔组织增生明显。结论膜引导组织再生术和自体静脉桥接法均可用于面神经缺损的修复,膜引导组织再生术的修复效果优于自体静脉桥接法。     

Dental regeneration and materials—a partnership

Considerable focus on the biocompatibility of dental materials over the last three decades has provided a platform for a wealth of studies on the cellular and molecular responses of the cells of the pulp to injury, both from the disease process and from subsequent restorative intervention. These studies have been fundamental to understanding not only how we can achieve a biocompatible response during restoration of dental disease but also how we can exploit the pulpal cellular responses to achieve wound healing and tissue regeneration in the dentine–pulp complex. This article examines the responses of the pulp to injury and the events leading to tissue regeneration. As new biologically based regenerative therapies emerge for the dental tissues, it is important that these develop in partnership with more traditional approaches using dental materials.