Microinjection of a growth factor cocktail affects activated microglia in the neocortex of adult rats

Microglia, as the resident immune cells in the central nervous system, play important roles in regulating neuronal processes, such as neural excitability, synaptic activity, and apoptotic cell clearance. Growth factors can activate multiple signaling pathways in central nervous system microglia and can regulate their immune effects, but whether growth factors can affect the morphological characteristics and ultrastructure of microglia has not been reported. After microinjecting 300 nL of a growth factor cocktail, including 10 μg/mL epidermal growth factor, 10 μg/mL basic fibroblast growth factor, 10 μg/mL hepatocyte growth factor and 10 μg/mL insulin-like growth factor into adult rat cortex, we found that the number of IBA1-positive microglia around the injection area increased significantly, indicating local activation of microglia. All CD68-positive labeling co-localized with IBA1 in microglia. Cell bodies and protrusions of CD68-positive cells were strongly attached to or were engulfing neurons. Characteristic huge phagosomes were observed in activated phagocytes by electron microscopy. The phagosomes generally included non-degraded neuronal protrusions and mitochondria, yet they contained no myelin membrane or remnants, which might indicate selective phagocytosis by the phagocytes. The remnant myelin sheath after phagocytosis still had regenerative ability and formed "myelin-like" structures around phagocytes. These results show that microinjection of a growth factor cocktail into the cerebral cortex of rodents can locally activate microglia and induce selective phagocytosis of neural structures by phagocytes. The study was approved by the Institute of Laboratory Animal Science, Beijing Institute of Basic Medical Sciences(approval No. IACUC-AMMS-2014-501) on June 30, 2014.     

间歇低氧对小鼠精子生成数量及活力的影响

目的：探讨间歇低氧对小鼠精子生成数量及活力的影响。方法将24只C57BL/6J雄性小鼠随机分为对照组和间歇低氧组，间歇低氧组置于自制的低氧舱中，舱内循环充入氮气和压缩空气，舱内氧浓度最高为（21±0.5）%、最低为（5±0.5）%，每次循环90 s，每天低氧8h；对照组置于与间断低氧舱相似的舱内，舱内循环充入洁净压缩空气。12周后将小鼠处死，测定各组小鼠的睾丸重、附睾重、精子数及精子活力。结果与对照组相比，间歇低氧组小鼠睾丸重、附睾重、精子生成的数量及活力均下降（P＜0.05）。结论间歇低氧能降低小鼠精子生成的数量及活力。     

Effects of High-Voltage Pulsed Radiofrequency on the Ultrastructure and Nav1.7 Level of the Dorsal Root Ganglion in Rats With Spared Nerve Injury

ObjectivesTo investigate the analgesic effect of high-voltage pulsed radiofrequency (HV-PRF) on the dorsal root ganglion (DRG) for neuropathic pain induced by spared nerve injury (SNI) in rats, especially the influence of this treatment on the DRG ultrastructure and voltage-gated sodium channel 1.7 (Nav1.7) level in the DRG.Materials and MethodsOne hundred fifty adult male Sprague-Dawley rats were randomly divided into five groups: Sham, SNI, Free-PRF, standard-voltage PRF (SV-PRF), and HV-PRF. The 45V-PRF and 85V-PRF procedures applied to the left L5 DRG were performed in SV-PRF group and the HV-PRF group, respectively, on day 7 after SNI, whereas no PRF was concurrently delivered in Free-PRF group. The paw mechanical withdrawal threshold (PMWT) was detected before SNI (baseline) and on days 1, 3, 7, 8, 10, 14, and 21. The changes of left L5 DRG ultrastructure were analyzed with transmission electron microscopy on days 14 and 21. The expression levels of Nav1.7 in left L5 DRG were detected by immunofluorescence and Western blot.ResultsCompared with the Free-PRF group, PMWT in the SV-PRF group and HV-PRF group were both significantly increased after PRF (all p < 0.05). Meanwhile, the PMWT was significantly higher in the HV-PRF group than that in the SV-PRF group on days 14 and 21 (all p < 0.05). There were statistically significant differences between the SV-PRF and Free-PRF groups (p < 0.05). Similarly, statistically significant difference was found between the HV-PRF and Free-PRF groups (p < 0.05). Especially, comparison of the SV-PRF group and the HV-PRF group revealed statistically significant difference (p < 0.05). The Nav1.7 levels were significantly downregulated in the SV-PRF group and HV-PRF groups compared to that in the Free-PRF group (all p < 0.01). A significantly lower Nav1.7 level was also found in the HV-PRF group compared to that in the SV-PRF group (p < 0.05).ConclusionsThe HV-PRF produces a better analgesic effect than SV-PRF applied to the DRG in SNI rats. The underlying mechanisms may be associated with improving the histopathological prognosis and the downregulation of Nav1.7 levels in the DRG.     

前庭器官超微结构研究及结构图构建

目的通过扫描电镜观察小鼠椭圆囊、球囊、壶腹超微结构,研究分析并据此构建前庭器官新的结构示意图。方法选取3个年龄段小白鼠各10只,分别是年轻组≤2个月、中年组2~12个月、老龄组>12个月。分离出椭圆囊、球囊、壶腹,采用扫描电镜技术样品制备方法制备样品,应用扫描电镜进行样品观察。结果扫描电镜下可以得到:①椭圆囊斑及球囊斑不同层面图片:表面为堆积并相互黏附的"表面耳石",表面耳石下是无结构胶状质;底层表面耳石深入到无结构胶质层里;无结构胶质层下面是毛细胞纤毛及"纤毛间耳石"层,不同纤毛束之间均有纤毛间耳石存在,立于支持细胞表面,表面平坦;蜂窝状胶质物质联接无结构胶质层、纤毛间耳石及毛细胞纤毛。②壶腹超微结构的图片:嵴帽是无结构的胶状质与壶腹外侧壁紧贴,但较易分离,嵴帽和壶腹外侧壁之间有纤细的晶状体物质,在壶腹嵴两侧壁上也有纤细晶体物质(壶腹嵴表面耳石);不同的毛细胞纤毛之间有耳石结构的存在(壶腹嵴纤毛间耳石)。结论通过对前庭器官扫描电镜的观察,发现了椭圆、球囊斑及壶腹的新结构成分,由此构建出新的前庭器官超微结构示意图。     

Consistent age-dependent declines in human semen quality: A systematic review and meta-analysis

Reduced fertility typically occurs among women in their late 30s, but increasing evidence indicates that advanced paternal age is associated with changes in reproduction as well. Numerous studies have investigated age-based declines in semen traits, but the impact of paternal age on semen parameter values remains inconclusive. Using data from 90 studies (93,839 subjects), we conducted a systematic review and meta-analysis to quantify the effect of male age on seven ejaculate traits (semen volume, sperm concentration, total sperm count, morphology, total motility, progressive motility and DNA fragmentation). Age-associated declines in semen volume, percentage motility, progressive motility, normal morphology and unfragmented cells were statistically significant and results generally seemed to be robust against confounding factors. Unexpectedly, sperm concentration did not decline with increasing male age, even though we found that sperm concentration declined over time. Our findings indicate that male age needs more recognition as a potential contributor to the negative pregnancy outcomes and reduced offspring health associated with delayed first reproduction. We suggest that greater focus on collection of DNA fragmentation and progressive motility in a clinical setting may lead to better patient outcomes during fertility treatments of aging couples.     

Structural adaption of axons during de‐ and remyelination in the Cuprizone mouse model

Multiple Sclerosis is an autoimmune disorder causing neurodegeneration mostly in young adults. Thereby, myelin is lost in the inflammatory lesions leaving unmyelinated axons at a high risk to degenerate. Oligodendrocyte precursor cells maintain their regenerative capacity into adulthood and are able to remyelinate axons if they are properly activated and differentiate. Neuronal activity influences the success of myelination indicating a close interplay between neurons and oligodendroglia. The myelination profile determines the distribution of voltage‐gated ion channels along the axon. Here, we analyze the distribution of the sodium channel subunit Nav1.6 and the ultrastructure of axons after cuprizone‐induced demyelination in transgenic mice expressing GFP in oligodendroglial cells. Using this mouse model, we found an increased number of GFP‐expressing oligodendroglial cells compared to untreated mice. Analyzing the axons, we found an increase in the number of nodes of Ranvier in mice that had received cuprizone. Furthermore, we found an enhanced portion of unmyelinated axons showing vesicles in the cytoplasm. These vesicles were labeled with VGlut1, indicating that they are involved in axonal signaling. Our results highlight the flexibility of axons towards changes in the glial compartment and depict the structural changes they undergo upon myelin removal. These findings might be considered if searching for new neuroprotective therapies that aim at blocking neuronal activity in order to avoid interfering with the process of remyelination.     

Evaluation of natural dyes for human spermatozoa morphology assessment

Dye extracts from plants have been valuable not only for the economy but also environmental sustainability. There have been many reports on the utilization of natural dyes extracted from various sources for staining of biological tissues. This study aimed to investigate the extraction of natural dye from black rice (Oryza Sativa), butterfly pea (Clitoria ternatea), fresh roselle (Hibiscus sabdariffa), and mulberry (Morus alba) to stain of human spermatozoa for morphology assessment. The results showed that black rice extracted from solvents C containing 5?ml of absolute ethanol, 10?g of potassium alum and 100?ml of distilled water is the best dye for human spermatozoa evaluation comparable to the rapid PAP and Dip quick® stain. Effectiveness of the process was found with black rice extract stain by using 2 steps for 15?min. There were no statistically significant differences in the parameters of head, midpiece, tail and background for human sperm morphology assessment comparable to rapid PAP and Dip quick® stain (p?>?0.05) unless the midpiece compartment when compare to rapid PAP. This finding suggests that the black rice extracted has potential for use as an alternative dye for human spermatozoa morphology evaluation. The usefulness of black rice extracts will decrease the expense for purchasing synthetic dyes and reduce their adverse effects on human and environment.     

ICSI联合IVM技术在卵巢低反应高龄患者中的应用

目的：探讨胞浆内单精子注射（ICSI）联合未成熟卵母细胞体外成熟（IVM）技术在卵巢低反应高龄患者中的应用价值。方法：回顾性分析2016年1月—2018年9月于郑州大学第一附属医院生殖医学中心行ICSI助孕治疗81个周期，年龄≥35岁，获卵数≤3个，且卵母细胞均未成熟。按促排卵方案分为微刺激方案组（n=37）和拮抗剂方案组（n=44），统计2组患者基础情况、实验室结果及妊娠结局。结果：①2组患者年龄、不孕时间、基础内分泌、基础窦卵泡数及抗苗勒管激素（AMH）比较，差异均无统计学意义（P＞0.05）；②拮抗剂方案组Gn总量较微刺激方案组高，差异有统计学意义（P＜0.05）；③微刺激方案组与拮抗剂方案组体外培养成熟率、2PN率、卵裂率、可利用胚胎率及优胚率比较差异均无统计学意义（P＞0.05）；④微刺激方案组与拮抗剂方案组的胚胎存活率、种植率和临床妊娠率比较差异无统计学意义（P＞0.05）。结论：对于卵巢低反应的高龄患者行ICSI联合IVM培养有利于提高卵母细胞成熟率及胚胎利用率，增加妊娠机会，微刺激方案Gn用量少对卵巢低反应患者是经济有效的选择。