CR16在特发性无精子症患者睾丸组织中的表达

目的:研究糖皮质激素区域表达基因16(corticosteroids and regional expression 16,CR16)在特发性无精子症患者睾丸组织中的表达,探讨CR16在精子发生中的作用。方法:采用免疫组化法和逆转录-聚合酶链反应(RT-PCR)检测CR16蛋白和CR16 mRNA在48例特发性无精子症患者(病例组)和10例正常生育男性(正常组)睾丸组织中的表达情况。结果:CR16蛋白表达于睾丸生精小管上皮,主要分布在支持细胞与生精细胞连接区,在特发性无精子症患者睾丸组织中的表达显著弱于正常生育男性睾丸组织中的表达。CR16 mRNA在病例组睾丸组织中的表达水平显著低于正常组。结论:特发性无精子症患者睾丸组织中CR16表达水平显著降低,这可能与无精子症相关。     

精子发生相关基因SPATA48在人和小鼠睾丸组织中的表达研究

目的通过分析精子发生相关基因SPATA48在人和小鼠睾丸组织中的表达,探讨SPATA48在精子发生中的作用。方法收集2010年1月至2017年12月在深圳市第二人民医院就诊的非梗阻性无精子症患者和正常生育者睾丸活检样本各60例;腹腔注射白消安方法建立小鼠无精子症模型,小鼠在不同时期处死后解剖留取各个脏器;提取小鼠各脏器组织、两组睾丸组织以及人精母细胞、精原细胞和支持细胞中mRNA;并购买人肺、肾、肝脏、脾脏等组织mRNA样本。利用RT-PCR方法检测SPATA48基因在各种细胞和组织器官、两组人睾丸组织中的表达水平,并利用免疫组化方法检测SPATA48蛋白在各种细胞和两组人睾丸组织中的表达水平,对各检测指标进行相关统计分析。结果 RT-PCR结果表明SPATA48基因特异表达在睾丸组织中,肺、肾脏、肝脏、脾脏等其他组织中均不表达;非梗阻性无精子症患者睾丸组织中无SPATA48基因表达,三种睾丸细胞仅精母细胞存在阳性表达。无精子症模型小鼠,随白消安处理时间的延长,SPATA48基因表达显著减弱(P0.05)。免疫组化结果显示,在正常生育者睾丸中SPATA48蛋白主要表达在精母细胞和圆形精子细胞中,在非梗阻性无精子症患者睾丸中无SPATA48蛋白表达。结论 SPATA48特异表达在人和小鼠睾丸组织中,无精子发生的睾丸组织无表达,提示SPATA48基因在精子发生中具有重要作用。     

异体异位发育小鼠睾丸组织中精子功能相关基因和蛋白的表达研究

目的:采用已建立的未成熟睾丸组织移植模型,研究异体异位生长发育的小鼠睾丸组织中精子功能相关基因Asrgl1,Gpx4,Asp,Prdx4和Spa17及其相关蛋白GPX-4和PRX-Ⅳ的表达,以迸一步评估该移植模型在生殖医学中应用的可行性.方法:以1～2 d小鼠睾丸组织为供体,7～12周雄性免疫缺陷小鼠为受体进行组织移植;移植8周后收取移植物组织,采用荧光定量PCR法检测移植物中Asrgl1,Opx4,Asp,Prdx4和Spa17的表达情况,并与正常小鼠睾丸组织进行对比;同时采用Western blot检测GPX-4和PRX-Ⅳ 2种蛋白的表达情况.结果:基因检测显示,5种受试基因在移植物中的表达与8周正常小鼠睾丸组织相比均有下调;蛋白分析提示,GPX-4和PRX-Ⅳ 2种蛋白在8周正常小鼠睾丸组织中表达而在小鼠睾丸组织移植物中表达缺陷.结论:几种主要在精子运动、精子顶体形成及精子的受精功能中发挥作用的基因和蛋白在移植物中表达下调及缺失提示,异体异位睾丸组织移植在临床应用的可行性还需进一步评估     

特发性不育患者外周血和睾丸组织中无精子因子基因表达的比较研究

目的 探讨男性特发性不育患者外周血和睾丸组织中无精子因子(AZF)基因表达的临床意义.方法 特发性不育患者62例,其中严重少精子症29例.无精子症33例.抽取患者外周血样本检测,8对引物为sY84和sY86(AZFa区).sY127和sY134(AZFb区),sY254和sY255(AZFc区)及内对照SRY(sY14)和ZFY.PCR检测包括MixA:SRY(sY14)-ZFY-sY84-sY134-sY255和MixB:SRY(sY14)-ZFY-sY86-sY127-sY254;穿刺获得患者睾丸标本,Trizol方法提取总RNA,反转录为cDNA.PCR检测包括SRY(sY14)-DFFRY-RBM-DAZ-β-actin.结果 外周血PCR结果显示:62例患者中AZF基因微缺失12例(19.4%),其中无精子症组9例,严重少精子症组3例.睾丸组织RT-PCR结果显示:62例均可见SRY阳性表达;RBM mRNA无表达2例,RBM和DAZ mRNA无表达1例,DAZ mRNA无表达12例,其中3例外周血细胞内DAZ基因正常.结论 特发性不育患者睾丸组织存在AZF基因表达缺失,睾丸组织RT-PCR检测有助于确定患者病因,结合外周血PCR检测有助于指导睾丸精子穿刺一胞浆内单精子注射治疗.     

IGF-1在非梗阻性无精子症睾丸组织中的表达

目的 研究胰岛素样生长因子-1(IGF-1)在非梗阻性无精子症睾丸组织中的表达并探讨其意义.方法 采用免疫组化EnVision法对32例非梗阻性无精子症人睾丸组织石蜡标本和13例人正常睾丸组织中的IGF-1的表达进行检测,通过统计学方法 分析其表达情况.结果 非梗阻性无精子症睾丸组织HE染色显示生精小管中无或仅少量精子,生精小管的生精上皮脱落、排列紊乱,生精小管壁部分有玻变萎缩.32例非梗阻性无精子症睾丸组织中IGF-1阳性表达4例,阴性表达28例,阳性率12.5%;13例正常睾丸组织中IGF-1阳性表达10例,阴性表达3例,阳性率76.9%;差异有显著性(P<0.01).结论 (1)IGF-1在睾丸组织中表达减少或缺失与生精功能障碍密切相关.(2)非梗阻性无精子症睾丸组织生精小管的生精上皮脱落、排列紊乱,生精阻滞.     

Fas和FasL 系统在非梗阻性无精子症睾丸中的表达

目的：为探讨睾丸生精功能障碍与细胞凋亡的关系，研究Fas和FasL系统在非梗阻性无精子症睾丸支持、间质和生精细胞中的表达。方法：对20例非梗阻性无精子症患者行睾丸开放性活检，常规病理检查，按Johnson评分法评价精子发生和发生障碍的程度；采用免疫组化SABC法对睾丸支持、间质和生精细胞进行Fas和FasL表达的检测。结果：睾丸活检生精功能评为8分有14例，3分有2例，6、5、4和2分各有1例。在20例非梗阻性无精子症睾丸的间质、支持和生精细胞均有Fas和FasI。的表达；而支持细胞Fas和FasL的阳性和强阳性表达率明显高于间质和生精细胞。结论：非梗阻性无精子症的睾丸支持、间质及生精细胞Fas和FasL的高表达与精子生成障碍是一致的，非梗阻性无精子症可能与生殖细胞过度凋亡密切相关。     

Survivin在睾丸生殖细胞瘤与无精子症患者睾丸组织中的表达

目的：探讨Survivin蛋白在睾丸生殖细胞瘤与无精子症患者睾丸组织中的表达情况及其临床意义。方法：采用免疫组织化学SABC法检测34例男性睾丸生殖细胞瘤和38例男性无精子症患者睾丸组织中Survivin蛋白的表达情况。结果：18例睾丸精原细胞瘤和16例非精原细胞瘤组织标本中Survivin蛋白的表达情况分别为88．9％和62．5％；在38例不同类型无精子症睾丸组织标本中，各组间Survivin蛋白的表达差异有统计学意义（P〈0．05），且表达情况与生精功能障碍严重程度呈负相关（r=-0．675，P〈0．05）。结论：在睾丸生殖细胞瘤和正常睾丸组织中可检测到Survivin蛋白的表达，Survivin蛋白可能是精子发生过程中的一个潜在标志物。     

Mm．158494基因在白消安引起的无精子症小鼠中的表达

目的建立一个小鼠无精子症模型,并探讨Mm.158494基因在无精子症中的表达及意义。方法建立无精子症小鼠模型,观察小鼠睾丸生精小管结构的变化。筛选与精子发生相关的睾丸特异性新基因,利用网络信息资源对该基因进行生物学信息分析,RT-PCR分析该基因在无精子症小鼠睾丸中的表达。结果生物信息学分析发现Mm.158494基因cDNA序列全长1046bp,含有762bp的完整开放阅读框。编码253个氨基酸、分子量为29.432kDa的蛋白质。小鼠无精子症模型中微弱表达Mm.158494基因。结论Mm.158494基因在无精子症中表达明显降低,提示该基因可能在精子发生中起重要作用。     

脐带间充质干细胞增强无精子症模型睾丸特异性基因表达

在生殖男科领域，探索人脐带间充质干细胞（HUCMSCs）对无精子症的治疗作用。HUCMSCs具有潜在的免疫抑制功能，并能够分泌多种细胞因子和生长因子，因此具有潜在的临床应用价值。作为探索，我们移植HUCMSCs进入无精子症小鼠睾丸间质，检测是否能够促进精子发生过程。从不同来源的脐带中分离HUCMSCs，移植进入白消安处理的小鼠睾丸间质中，采用注射生理盐水和HEK293细胞作为对照，对侧睾丸不注射。三周之后，RT-PCR检测10个生殖细胞特异性表达的基因，并检N3个特异性蛋白的表达。结果表明，注射人脐带间充质干细胞之后的表达明显高于对照组，证实生殖细胞特异性基因的表达上调，从而表明HUCMSCs对睾丸生精功能的恢复具有促进作用，为治疗无精子症探索一条新的途径。     

无精子症和生精功能正常者睾丸组织中血红素加氧酶的表达与差异性研究

目的:对人类睾丸组织中表达血红素加氧酶(HO)的细胞进行定位;通过测定原发性无精子症及梗阻性无精子症患者睾丸组织中血红素加氧酶1(HO-1)的表达量与正常睾丸组织中HO-1表达量的差异性,来探讨其与无精子症发病的相关性。方法:应用免疫组化方法对人类睾丸组织中表达HO的细胞进行定位;采用逆转录-荧光定量PCR(FQ-PCR)方法定量检测无精子症患者与正常人睾丸组织HO-1及HO-2基因水平的表达量;应用W est-ern印迹检测各组之间HO蛋白水平表达量。结果:在正常睾丸组织,HO-1主要表达在支持细胞上;而HO-2在支持细胞和各级生精细胞中均有表达;FQ-PCR结果显示非梗阻性无精子症患者睾丸组织HO-1、HO-2的表达量均显著低于正常组及梗阻性无精子症组(P<0.05),差异具有统计学意义。而梗阻性无精子症患者睾丸组织表达HO-1、HO-2的量与正常组相比无显著性差异。W estern印迹结果显示HO-1蛋白水平的表达量差异与基因水平一致。而HO-2的蛋白水平在各组之间表达没有显著性差异。结论:非梗阻性无精子症患者睾丸组织中HO的表达量显著性降低,且HO-1无论是蛋白水平还是基因水平的差异一致。HO-1可以通过抗炎、抗氧化、抗凋亡的机制保护睾丸组织免受各种应激的损伤,从而维护正常的生精功能。可见,HO-1的减少可能与生精功能低下相关,这可能是非梗阻性无精子症的发病机制之一。     

Histochemical and contractile properties of striated muscles of urethra and levator ani of dogs and sheep

AIMS: To understand their possible importance in long- and short-term control of continence, some properties of the striated muscles of the urethra and pelvic floor (levator ani) of dogs and sheep were investigated, especially fiber types and contractile characteristics. MATERIALS AND METHODS: Striated muscles of urethra and levator ani of 29 male and 6 female dogs and 11 male and 6 female sheep were removed and cut into strips. Some strips were frozen and stained for ATPase at pH 9.4 and 4.3 for fiber typing; others were set up in an organ bath to study contractile responses to nerve stimulation. RESULTS: All muscles contained both type I (slow) and type II fibers, ranging from 97% type II in female greyhound urethra to 60% in female sheep levator ani. For each muscle, there were fewer type II muscles in sheep than in dog. The diameters of the urethral fibers were about 60% of the levator ani in dogs and 34% in sheep. Contraction of the urethral muscle was faster than for levator ani and declined to about 80% of the peak, 500 msec after the beginning of stimulation at 20 Hz. The levator ani contraction rose to a steady level as long as stimulation continued. CONCLUSIONS: Both the levator ani and urethral striated muscles contain slow and fast fiber types. The levator ani muscles are capable of sustained contraction with rapid onset which will produce long-term closure of the urethra. The circular urethral muscle contraction was faster but less well maintained.     

Incorporation and release of85Sr and14C-proline-hydroxyproline in mineral and collagen of bone and incisor teeth of growing rats

The extent to which exchange and reutilization processes of mineral tracers affect skeletal mineral accretion and resorption measurements was evaluated by comparing the rates of appearance and disappearance of⁸⁵Sr and¹⁴C-proline-hydroxyproline in bones and teeth in growing rats for 12 days following simultaneous parenteral injection of these tracers. Expressions for the relative rates of collagen synthesis and breakdown, which unlike mineral metabolism are considered not to be complicated by exchange phenomena, were based on¹⁴C-proline conversion to¹⁴C-hydroxyproline; the specific activity of the latter was determined. Both the mineral and the collagen specific activities reflected the rates and patterns of growth of the samples assayed; rapid growth and a short interval of time between formation and resorption of tissue in themetaphyseal bone which contains the cartilagineous growth plate, slow growth and an interval of time between formation and resorption of tissue indiaphyseal bone and incisor teeth which is longer than the 12 days of the experiment. However, in metaphyseal bone the specific activity collagen/mineral ratio dropped by one half during the 4–12 day interval in contrast to diaphyseal bone and incisor teeth in which no change in this ratio was observed during this period of time. The data indicate that collagen in the metaphyseal growth zone is removed by resorption before it has become fully mineralized, and that exchange is a relatively unimportant factor in the long term kinetics of bone mineral.

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Zusammenfassung Das Ausmaß, bis zu welchem Austausch- und Wiederverwendungsprozesse der mineralen Tracer die Messungen des mineralen Skelett-Auf- und Abbaues beeinflussen können, wurde ausgewertet; zu diesem Zweck wurde die Geschwindigkeit des Auftretens und Verschwindens von⁸⁵Sr und von¹⁴C-Prolin-Hydroxyprolin in Knochen und Zähnen von wachsenden Ratten während der 12 auf die simultane parenterale Injektion dieser Tracer folgenden Tage verglichen.Der Ausdruck für die relative Geschwindigkeit des Kollagen-Auf- und Abbaues, bei welchem im Gegensatz zum Mineralmetabolismus kein Mitwirken des Austauschphänomens vermutet wird, basiert auf der Umwandlung von¹⁴C-Prolin zu¹⁴C-Hydroxyprolin; die spezifische Aktivität des letzteren wurde bestimmt.Aus der spezifischen Aktivität des Minerals sowie jener des Kollagens konnten die Geschwindigkeit und die Art des Wachstums der untersuchten Proben ersehen werden, d.h.schnelles Wachstum und ein kurzes Zeitintervall zwischen Bildung und Resorption des Gewebes imKnochen der Metaphyse, die auch die knorpelige Wachstumsplatte enthält, und andererseitslangsames Wachstum und längeres Zeitintervall (länger als die 12 Tage des Experimentes) zwischen Bildung und Resorption des Gewebes imKnochen der Diaphyse und in den Schneidezähnen. Immerhin fiel die spezifische Aktivität des Kollagen/Mineral-Anteils im Knochen der Metaphyse während dem 4–12tägigen Zeitintervall auf die Hälfte, im Gegensatz zum Knochen der Diaphyse und der Schneidezähne, bei welchen während dieser Zeitspanne kein Unterschied in diesem Verhältnis beobachtet wurde.Diese Ergebnisse zeigen, daß Kollagen in der Wachstumszone der Metaphyse durch Resorption verschwindet, bevor es ganz mineralisiert ist, und daß der Austausch ein relativ unwichtiger Faktor in der Kinetik auf lange Sicht des Knochenminerals ist.

     

Instrumented ligamentotaxis and stabilization of compression and burst fractures of dorsolumbar and mid-lumbar spines

Background:

Controversy continues regarding the best treatment for compression and burst fractures. The axial distraction reduction utilizing the technique employing the long straight rod or curved short rod without derotation to reduce fracture are practised together with short segment posterolateral fusion (PLF). Effects of the early postoperative mobilization without posterolateral fusion on reduction maintenance and fracture consolidation were not evaluated so far. The present prospective study is designed to assess the effectiveness of i) reduction and restoration of sagittal alignment, ii) no posterolateral fusion on the reduced, fractured vertebral body and injured disc, iii) fracture consolidation and iv) the fate of the unfused cephalad and caudal injured motion segments of the fractured vertebra.

Materials and Methods:

The study includes 15 Denis burst and two Denis type D compression fractures between T₁₂ and L₃. The lordotic distraction technique was used for ligamentotaxis utilizing the contoured short rods and pedicle screw fixator. Three vertebrae including the fractured one were fixed. The patients after surgery were braced for ten weeks with activity restriction for 2-4 weeks. The patients were evaluated for change in vertebral body height, sagittal curve, reduction of retropulsion, improvement in neural deficit. The unfused motion segments, residual postoperative pain and bone and metal failure were also evaluated.

Results:

The preoperative and postreduction percentile vertebral heights at, zero (immediate postoperative), at three, six and 12 months followup were 62.4, 94.8, 94.6, 94.5 and 94.5%, respectively. The percentages of the intracanal fragment retropulsion at preoperative, and postoperative at zero, 3, 6 and 12 months followup were 59.0, 36.2,, 36.0, 32.3, and 13.6% respectively.The preoperative and postreduction percentile loss of the canal dimension and at zero, three, six and 12 months were 52.1, 45.0, 44.0, 41.0 and 29% respectively suggesting that the under-reduced fragment was being resorbed gradually by a remodeling process. The mean initial kyphosis of 33° became mean 2° immediately after reduction and mean 3° at the final followup. The fractured vertebral bodies consolidated in an average period of ten weeks (range 8-14 weeks). The restored disc heights were relatively well maintained throughout the observation period. All paraparetic patients recovered neurologically. There were no postoperative complications.

Conclusion:

Instrument-aided ligamentotaxis for compression and burst fractures utilizing the short contoured rod derotation technique and the instrumented stabilization of the fractured spine are found to be effective procedures which contribute to the fractured vertebral body consolidation without recollapse and maintain the motion segment function.     

Principles and Practice of Hemofiltration and Hemodiafiltration

There is growing interest in the convective dialysis therapies, hemofiltration (HF) and hemodiafiltration (HDF). Both require dialysis membranes which are highly permeable to solutes as well as fluid, and in both cases large volumes of ultrafiltration are the condition for convective transport. In HDF the convection is combined with diffusion, and as a consequence, maximum clearance over the entire molecular weight spectrum is achieved. Optimal forms of HDF provide urea clearance 10–15% higher than the corresponding diffusive mode. The larger the solute, the greater is the impact of convection, and β₂-microglobulin (β₂m) levels may be up to 70% reduced. Traditional postdilution HF provides high clearance of medium sized and large molecules. Satisfactory clearance of small solutes requires blood flows in excess of 500 ml/min. With access to practically unlimited volumes of substitution solution through on-line ultrafiltration, predilution HF can now be used. This increases the clearance of small solutes to an acceptable range. For HDF as well as HF, large patient populations consistently treated for longer periods of time are needed to make valid outcome comparisons with other therapies.     

骨折不愈合与延迟愈合的成因与治疗

目的探讨骨折不愈合与延迟愈合的成因、报肯治疗的方法与设果。方法对1990年7月～2004年12月间收治的107例骨折不愈台、54例骨折延迟愈合2例先天性胫骨骨不连进行回顾性研究，分析原因，随访治疗结果。18例延迟愈合行保守治疗，本组其他145例行手术治疗，结果除2例先天性胫骨骨不连外，其余161例的成因中均有医源性因素。10例失去随访，153例平均随访17（6-28）个月，骨折均获骨性连接，愈合时间平均10（6-14）个月，肢体功能恢复良好，结论医源性技术缺陷是骨折不愈合与延迟愈合的主要原因，针对各种不同因素进行合理治疗可获得满意效果。     

Recognition and management of phaeochromocytoma and paraganglioma

Phaeochromocytomas and paragangliomas (PPGL) are catecholamine-secreting neuroendocrine tumours arising from the chromaffin cells in the adrenal medulla. These tumours may be identified incidentally, as part of a work-up for multiple endocrine neoplasia or following haemodynamic surges during unrelated procedures. Advances in perioperative management and improved management of intraoperative haemodynamic instability have significantly reduced surgical mortality from around 40% to less than 3%. Surgery is the definitive treatment in most cases and laparoscopic resection where possible is associated with improved outcomes. Anaesthetic management of PPGL cases represents a unique haemodynamic challenge both before and after tumour resection. In this article we describe the physiology of these tumours, their diagnosis, preoperative optimization methods, intraoperative anaesthetic management and management of postoperative complications.     

Physiology and pharmacology of nausea and vomiting

Nausea and vomiting are both very unpleasant experiences. The physiology is poorly understood; however, understanding what we do know is key to tailoring a preventative or therapeutic antiemetic regime. There are two key sites in the central nervous system implicated in the organization of the vomiting reflex: the vomiting centre and the chemoreceptor trigger zone. There are five key neurotransmitters involved in afferent feedback to these areas. These are histamine (H₁ receptors), dopamine (D₂), serotonin (5-HT₃), acetyl choline (muscarinic) and neurokinin (substance P). Postoperative nausea and vomiting will occur in around one-third of elective patients who have no prophylaxis. This can result in many detrimental effects including patient dissatisfaction, unplanned admission and prolonged recovery. It is therefore essential that clinicians understand how they can prevent and treat nausea and vomiting using either a single agent or a combination of antiemetics to target relevant receptors. Commonly used drugs include antihistamines, dopamine antagonists, serotonin antagonists and steroids. More novel agents are being developed such as aprepitant, a neurokinin receptor antagonist, palonosetron, a 5HT₃ receptor antagonist and nabilone, a synthetic cannabinoid.     

Physiology and pharmacology of nausea and vomiting

Nausea and vomiting are both very unpleasant experiences. The physiology is poorly understood; however, understanding what we do know is key to tailoring a preventative or therapeutic antiemetic regime. There are two key sites in the central nervous system implicated in the organization of the vomiting reflex: the vomiting centre and the chemoreceptor trigger zone. There are five key neurotransmitters involved in afferent feedback to these areas. These are histamine (H₁ receptors), dopamine (D₂), serotonin (5-HT₃), acetyl choline (muscarinic) and neurokinin (substance P). Postoperative nausea and vomiting will occur in around one-third of elective patients who have no prophylaxis. This can result in many detrimental effects including patient dissatisfaction, unplanned admission and prolonged recovery. It is therefore essential that clinicians understand how they can prevent and treat nausea and vomiting using either a single agent or a combination of antiemetics to target relevant receptors. Commonly used drugs include antihistamines, dopamine antagonists, serotonin antagonists and steroids. More novel agents are being developed such as aprepitant, a neurokinin receptor antagonist, palonosetron, a 5HT₃ receptor antagonist, and nabilone, a synthetic cannabinoid.