Andrology: Factors influencing the outcome of in-vitro fertilization with epididymal spermatozoa in irreversible obstructive azoospermia

Microsurgical epididymal sperm aspiration (MESA) and in-vitrofertilization (IVF) were found to offer limited opportunityfor fatherhood to 45 men with obstructive azoospermia, due principallyto poor embryo implantation. Adequate sperm preparations wereobtained in 46/50 treatment cycles (92%), with the best motilityfound in the caput epididymis in 89% of cases. The mean fertilizationrate was 11.2% and fertilization occurred in 23 cycles (50%),with embryo transfer arising from 12/26 men with was aplasia(CAV), 4/9 with genital tract obstruction (EV) and 7/11 withirreversible vasectomy (VV). The overall implantation rate waslow, 8.7% per embryo transfer (11.7% per 2-3 embryo transfers)and was not improved by Fallopian transfer. There were two pregnancies(4% per cycle), both in the EV group where embryo formationand implatation (2/4, 50% per cycle) were optimum even thoughsperm preparations were paradoxically inferior to the CAV andVV groups. The spermatozoa retrieved in the two successful EVcycles were appreciably blood contaminated. Analysis of the21 failed embryo transfers showed delayed fertilization in 10cycles, cystic fibrosis (CF) mutation or familial disease in7/12 CAV men and the VV men were older (P<0.001). A pregnancywhich miscarried arose from a case of Young's syndrome, a carrierof CF mutation DF508. Male factors could thus be implicatedin the high embryo wastage of MESA cycles and might also beinfluencing implantation in other IVF procedures. Where feasible,male reconstructive surgery is preferable unless fertilizationcan be improved, possibly by speedier retrieval techniques orby permitting sperm capacitation in vitro, but probably moreeffectively by micro-assisted insemination.     

HLA-DR and H-2E transgenes differentially mediate TCR-specific positive selection

The use of HLA transgenic mice in models of immunity and diseaseassumes that human MHC molecules are able to contribute towardthe positive selection of the mouse TCR repertoire. As an initialstep towards analysis of this we have compared the relativeability of DR/Eß or E/Eß complexes to induceT cell receptor (TCR) positive selection in H-2Ea and HLA-DRAtransgenic mice lacking endogenous E. The results show that,like E/Eß, the hybrid DR/ß complexes arecapable of mediating positive selection of V_ß2⁺;,V_ß6⁺, and V_ß10⁺ cells. However, differenceswere found between the effects of the two transgenes. Thus,while V_ß6⁺ cells were efficiently selected in bothH-2Ea and DRA transgenic mice, positive selection of V_ß10⁺cells was less apparent in the DRA transgenic mice. Variationbetween Ea and DRA transgenic mice is consistent with the notionthat this process is dependent on differential binding of endogenouspeptides to the E/Eß and DR/Eß complexes.Furthermore, contrary to expectations, in neither set of micewas positive selection limited solely to the CD4⁺ subset. Thus,examples were found in which V_ß-specific positiveselection was confined to either the CD4⁺ or CD8⁺ subsets, andothers in which both subpopulations were concomltantly increased.In the case of V_ß2 positive selection, H-2Ea transgenicmice showed expansion of these cells in both the CD4⁺ and CD8⁺subpopulations whlle in DRA transgenic mice this occurred predominantlyin the CD8⁺ subpopulatlon.     

Signal transduction in human B cells initiated via Ig{beta} ligation

Ig and Igß heterodimers are non-covalently associatedwith Ig to compose the antigen receptor complexes on B cells.The demonstration that different sets of tyrosine kinases bindto the cytoplasmic tails of Ig and Igß suggests thatIg and Igß may activate distinct second messengerpathways. In this study, we examined the effects of mAbs againstan exposed epitope of human Igß on pre-B and B celltriggering. Cross-linkage of Igß on B cells leadsto activation of tyrosine kinases, hydrolysis of phosphatidylinositides,and elevation of intracellular Ca²⁺, effects qualitatively identicalto those of anti-_µ mAbs. Our observations thus indicatethat cross-linking of Igß does not segregate signaltransduction pathways connected with the cytoplasmic talls ofIg and Igß. Ig ligation has been reported to be moreeffective in triggering pre-B than B cells, whereas our resultsindicated that Igß ligation is more efficient in triggeringB than pre-B cells. In addition to their activation properties,the anti-Igß mAbs effectively modulated B cell receptorcomplexes and blocked terminal differentiation of all plasmacell isotypes. The findings support the idea that anti-Igßcould serve as a universal B cell immunosuppressant.     

Fertility of ejaculated and testicular megalohead spermatozoa with intracytoplasmic sperm injection

In this study the fertility and outcome of intracytoplasmic sperm injection (ICSI) using megalohead spermatozoa from the ejaculates and testicles was evaluated. Seventeen males with megalohead and pinhead sperm forms in their ejaculate were studied in 22 cycles. A high number of sperm heads without tails and abundant round spermatid forms were commonly observed. Round-headed spermatozoa were seldom accompanied by these severely abnormal spermatozoa. The majority of megalohead spermatozoa were observed to have multiple tails, were predominant in the sample, and were used for ICSI. Ejaculated megalohead spermatozoa were used for ICSI in 15 cycles, while testicular spermatozoa were used in seven cycles where there were no vital spermatozoa or spermatozoa of low vitality in the ejaculate. The same abnormal morphology was observed in the testicles as in the ejaculated spermatozoa in the same males. Mean (+/- SD) low motility 4.7 +/- 5.6% and sperm count (3.8 +/- 4.19 x 10(6)) were common findings in these severely teratozoospermic patients. A low fertilization rate (43.2%) was achieved by using megalohead sperm forms (group I, n = 17) in comparison with the control group (60.2%) which had zero normal sperm morphology according to strict criteria (group II, n = 30) (P <0.01). Furthermore, a low pregnancy rate (9.1%) was obtained in the megalohead sperm group in comparison with the control group (40%) (P <0.05). Low fertilization and pregnancy rates may be due to a high incidence of chromosomal abnormalities from severely defective spermatozoa in the ejaculate. Couples should be counselled and warned about possible low fertilization and pregnancy rates with ICSI when only pinhead and megalohead forms with a high number of sperm heads without tails are present in the ejaculate.     

Early effects of aldosterone on the basolateral potassium conductance of A6 cells

Summary Aldosterone increases the basolateral conductance in target epithelia. The basolateral membrane of tight epithelia contains two different types of K⁺ conductances (G_K), a resting and a volume-activated G_K. We have studied the early effects (at 4 hours) of 500 nmol/l aldosterone on the basolateral membrane G_k of A6 cells (a Xenopus laevis kidney cell line), after the permeabilization of the apical membrane with amphotericin B. In the presence of a 97 to 3 mmol/l apical to basolateral K⁺ gradient, the resting, inward rectifying G_K was similar in control and aldosterone treated cells. In contrast, aldosterone induced a 2-fold increase of the volume-activated quinidine sensitive G_K.     

Suppression of human lymphocyte responses to specific and non-specific stimuli in human onchocerciasis.

Characterization of in vitro lymphocyte responsiveness was performed on selected groups of onchocerciasis patients from Sudan and Sierra Leone. These patients manifested a very broad range of clinical signs and showed widely divergent parasite infection intensities. Lymphocyte proliferative responses to soluble Onchocerca volvulus antigen (sAg) were poor in infected persons; mitogen and PPD responses were maintained in the normal range in one group of patients from southwestern Sudan, but were profoundly depressed in a group from N.E. Sudan. Proliferative responses and interferon-gamma (INF-gamma) secretion were very significantly depressed in the presence of live microfilariae of O. volvulus or secretions/excretions (S/E) from microfilariae (mf) or from female, but not male, adult parasites. Lymphocyte responses were maintained near normal when exogenous IL-2 was added to these cultures. The results indicate that O. volvulus infection and its clinical consequences are not consistently associated with systemic deficits in immune responsiveness. However, suppression of lymphocyte reactivity by mf and S/E in vitro suggests that direct parasite intervention in host cell responses could be taking place in vivo, perhaps at the local microenvironment level; mediated by effects on cytokine production.     

Changes in anti-Müllerian hormone serum concentrations over time suggest delayed ovarian ageing in normogonadotrophic anovulatory infertility

BACKGROUND: Anti-Müllerian hormone (AMH), produced by growing pre-antral and early antral ovarian follicles, has been shown to be a useful marker for ovarian ageing. Serum AMH concentrations are elevated during reproductive life in anovulatory women, especially in those patients exhibiting polycystic ovaries (PCO). The current study was designed to investigate whether the decrease in AMH serum concentrations over time is different comparing women with normogonadotrophic anovulation [World Health Organization (WHO) group 2 (including polycystic ovary syndrome (PCOS)] and normo-ovulatory controls. METHODS AND RESULTS: AMH serum levels were assessed on two occasions in 98 patients suffering from WHO 2 anovulatory infertility as well as in 41 normo-ovulatory premenopausal women. Median time interval between both visits was 2.6 years (range 0.3-9.0) for WHO 2 patients compared with 1.6 years (range 1.0-7.3) in controls. Serum AMH concentrations were significantly (P < 0.0001) elevated on both occasions in WHO 2 patients (AMH1, median = 7.5 microg/l, range 0.1-35.8; and AMH2, median = 6.7 microg/l, range 0.0-30.6) compared with controls (AMH1, median = 2.1 microg/l, range 0.1-7.4; and AMH2, median = 1.3 microg/l, range 0.0-5.0). Regression analysis, corrected for age, indicated a significant relative decrease in serum AMH concentrations over time for both groups (P < 0.001). However, the decline in serum AMH in WHO 2 patients was significantly less compared with controls (P = 0.03). CONCLUSION: The present longitudinal study shows that serum AMH concentrations decrease over time both in women presenting with WHO 2 anovulatory infertility and in normo-ovulatory controls. The decrease in WHO 2 patients is less pronounced despite distinctly elevated concentrations. This observation may suggest retarded ovarian ageing and hence a sustained reproductive life span in these patients.     

凋亡抑制蛋白XIAP和促凋亡因子Smac在胰腺癌化疗抵抗中的作用

目的探讨x-相关凋亡抑制蛋白（XIAP）和促凋亡因子Smac在胰腺癌细胞化疗抵抗中的作用，以及转染胞浆表达型Smac基因靶向下凋XIAP对化疗药物诱导的胰腺癌细胞凋亡的影响。方法应用流式细胞术检测顺铂、5-FU介导的Panc-1、BXPC-3的凋亡率及胞浆染色分析细胞XIAP表达变化，Western blot分析XIAP、Smac、Caspase-3表达水平；构建pEGFP-NI／Smac真核表达载体并转染胰腺癌Panc-1细胞，流式细胞术检测转染Smac基因前后Panc-1细胞的凋亡敏感性。结果与BXPC-3细胞相比，Panc-1对顺铂或5-FU介导的凋亡具有较强抵抗性，Western blot分析显示Panc-1细胞高表达XIAP，在化疗药物作用下化疗敏感细胞BXPC-3胞浆内XIAP水平下降明显多于Panc-1细胞，而且凋亡的BXPC-3细胞释放入胞浆内的成熟Smac蛋白水平明显高于Panc-1细胞。转染胞浆表达型Smac基因至化疗抵抗Panc-1细胞，可明显下调其XIAP表达水平，促进效应Caspase-3分子活化，显著提高顺铂、5-FU诱导的细胞凋亡率。结论胰腺癌细胞XIAP的表达水平下调与其化疗敏感性有关，XIAP是克服化疗抵抗的重要靶分子，而上调Smac活性蛋白的胞浆表达作为一种有效调节信号，通过拮抗XIAP的凋亡抑制作用协同化疗药物促进胰腺癌细胞凋亡。