神经移位治疗火器性肌皮神经和腋神经损伤

在观察和测量100个上肢解剖标本的基础上,我们设计了带血管的胸背神经和下肩胛下神经移位,一期修复一例火器性肌皮神经和腋神经损伤.术后8个月复查功能恢复甚为满意.     

Wg3h细胞系转染PSV_2-neo质粒前后生长特性及表面超微结构改变的研究

本文对转染PSV_2-neo质粒后的Wg3h细胞系(Wg3h-neo)在长期传代中的生长特性和表面超微结构与母系Wg3h细胞进行了比较研究。结果发现:转染后Wg3h细胞的DNA合成及生长速度明显高于母系Wg3h细胞,生长饱和密度增大。在软琼脂培养中不形成集落,接种在裸鼠不长肿瘤。在扫描电镜下,细胞表面的微绒毛较母系Wg3h细胞丰富。Southern印迹杂交实验证明PSV_2-neo质粒已整合到宿主细胞的基因组中。转染后Wg3h细胞的生长特性和表面超微结构均发生了某些转化特征。     

Regulatory role of mature B cells in a murine model of inflammatory bowel disease

The spontaneous chronic colitis in TCR alpha mutant (TCRalpha(-/-)) mice mediated by CD4(+) TCRalpha(-)beta(+) T cells is more severe in the absence of mature B cells, suggesting a suppressive role of B cells and Ig in the development of chronic colitis. To investigate the direct role of B cells in the suppression of this colitis, cell transfer studies were performed in TCRalpha(-/-) x Igmu(-/-) (alphamu(-/-)) double-knockout mice. The chronic colitis was markedly attenuated in alphamu(-/-) mice after the adoptive transfer of peripheral B cells from TCRalpha(-/-) mice into 3- to 4-week-old alphamu(-/-) mice prior to the development of colitis. Furthermore, transfer of mature B cells from TCRalpha(-/-) mice markedly decreased the number of pathogenic colonic CD4(+) TCRalpha(-)beta(+) T cells in alphamu(-/-) mice with established colitis. This B cell effect required the presence of functional co-stimulatory molecules CD40 and B7-2 (CD86) but not B7-1 (CD80). These results indicate that mature B cells play an important role in the development of chronic colitis in TCRalpha(-/-) mice by directly regulating the pathogenic T cells (CD4(+) TCRalpha(-)beta(+) T cells).     

Lipofectin介导转移的外源性ADA基因在反复上感儿T淋巴细胞的表达

本文对４１例健康儿童和１７例反复上呼吸道感染患儿外周血淋巴细胞腺苷脱氨酶（ＡＤＡ）活性进行了检测。在此基础上筛选出２例反复上感伴ＡＤＡ活性低下患儿。在体外对这２例患儿的外周血Ｔ淋巴细胞进行培养后，以Ｌｉｐｏｆｅｃｔｉｎ（脂质体）介导的方法对其进行了外源性ＡＤＡ基因的基因转移。结果显示：２例患儿体外培养淋巴细胞ＡＤＡ活性较转基因前升高。同步进行的标志基因ｐＢＬａｃＺ的基因转移的检测结果也直观地证实了Ｌｉｐｏｆｅｃｔｉｎ介导的基因转移是成功的。该研究为ＡＤＡ－ＳＣＩＤ淋巴细胞基因治疗的研究提供了初步的体外实验资料。     

Impact of elective single embryo transfer on the twin pregnancy rate

BACKGROUND: It is unclear how the implementation of elective single embryo transfer in clinical practice would affect clinical pregnancy and delivery rates and multiple birth rates. METHODS: This retrospective study analysed 1871 IVF/ICSI cycles carried out from 1997 to 2001 in the IVF programme of a single university infertility clinic. RESULTS: The number of elective single embryo transfers increased from 11 to 56%. At the same time the clinical pregnancy rate was relatively stable; mean 34.0% (range 28-42). The number of embryos per embryo transfer decreased from 1.8 to 1.3. The multiple pregnancy and delivery rates dropped markedly from 25 to 7.5% and from 25 to 5% respectively. CONCLUSIONS: An elective single embryo transfer programme can be adopted in daily practice that decreases the twinning rate to <10% and does not affect the overall pregnancy rate.     

Morphological evaluation of human embryos and derivation of an embryo quality scoring system specific for day 3 embryos: a preliminary study

A scoring system specific for day 3 embryos has not been extensively explored. Most IVF laboratories continue to grade embryos solely on the basis of cell number and percentage fragmentation as was traditionally done for day 2 embryos. Additional morphological features, some unique to day 3 embryos, may be useful in selecting embryos most likely to blastulate and implant. The objective of this study was to derive an embryo scoring system for day 3 transfers which is predictive of positive pregnancy outcomes. A total of 316 transferred embryos from 93 patients was recorded on videotape and evaluated. The following parameters were used to grade the embryos: cell number, fragmentation pattern (FP), cytoplasmic pitting, compaction, equal sized blastomeres, blastomere expansion and absence of vacuoles. The clinical pregnancy rate was 41.9%, with an implantation rate of 18% per embryo transferred. The mean number of embryos transferred per patient was 3.4. Three formulae were derived to score embryo quality in each transfer based on the average score of individual embryos transferred. In the first scoring system, cell number alone was used to predict pregnancy outcome. The second scoring system was based on blastomere number and the observed FP. The third scoring system utilized both blastomere number and FP but also combined this with five morphological criteria to yield a final day 3 embryo quality (D3EQ) score. We found the D3EQ score to be prognostic of pregnancy outcome. This study suggests that although cell number and FP are certainly predictors of positive pregnancy outcomes, additional parameters specific to day 3 embryos should be used to stratify a cohort of embryos further.     

Specific cytotoxic T lymphocytes in gene therapy

 Cytotoxic T lymphocytes possess the capacity to lyse target cells which express antigens on their surface recognized by the T cell receptor. These cells are crucial in the body’s defense against foreign antigens. It has long been a goal of tumor biology to utilize T cells, specialized in the elimination of unwanted cells, for the treatment of cancer. The killing activity of T lymphocytes is restricted to specific antigen-presenting cells. For this reason the use of cytotoxic T cells in the elimination of cancer cells is limited to cancer cells which present neoantigens on their surface. To circumvent this limitation we describe a procedure in which the ζ component of the T cell receptor is genetically manipulated and equipped with an extracellular recognition domain. Introduction of a chimeric gene, consisting of the ζ chain of the T cell receptor and a single-chain antibody domain, into cytotoxic T lymphocytes results in T cells with a predetermined recognition specificity for particular tumor cells. The MHC restriction of target cell recognition can be avoided and tumor cells recognized by the single chain antibody domain can be recognized and lysed. Retroviral-mediated gene transduction was used to introduce chimeric ζ chain constructs into primary T cells of mice. The cocultivation of retrovirus producing helper cells with in vitro activated T lymphocytes led to a high gene transduction efficiency into primary T cells. These primary T cells assumed a predetermined specificity for target cell recognition and lysis. The production and provision of tumor cell specific T lymphocytes might not be sufficient to eradicate large tumors in vivo. Using a Schwannoma cell line, we showed that transplanted tumors secrete transforming growth factor β and thereby stifle the action of lymphocytes. We suggest that a coordinated strategy including the suppression of tumor cells specific antilymphocyte action and the provision of tumor cell specific T cells might be required to successfully eliminate tumor cells in vivo. Received: 13 September 1996 / Accepted: 30 October 1996     

Elective single day 3 embryo transfer halves the twinning rate without decrease in the ongoing pregnancy rate of an IVF/ICSI programme

BACKGROUND: Data on the effect of elective single embryo transfer (eSET) on the total and multiple pregnancy rates of an IVF/ICSI programme are reported. METHODS AND RESULTS: A retrospective cohort analysis of eSET was carried out over a 4 year period. A total of 1559 cycles resulted in 1464 transfers; 299 transfers of one top quality embryo (20.4%) and 86 of one non-top quality embryo (5.9%) yielded 149 conceptions (49.8%) with 105 ongoing pregnancies (35.1%) and 26 conceptions (30.2%) with 19 ongoing implantations (22.1%) respectively; 1079 transfers of two (n = 853; 58.3%) or more than two (n = 226; 15.4%) embryos yielded 366 ongoing pregnancies (33.9%). The ongoing pregnancy rates for the years between 1998 and 2001 were 35.9, 27.9, 31.9 and 31.0% per oocyte retrieval and 38.5, 29.4, 34.1 and 33.2% per transfer. There were no differences in pregnancy rates between any of the years. The average ongoing pregnancy rate (>12 weeks) over the 4 years was 31.5% per started cycle and 33.5% per transfer; the average number of embryos transferred decreased from 2.26 (1998) to 1.79 (2001); the multiple pregnancy and twinning rates dropped from 33.6 and 29.5% (1998) to 18.6 and 16.3% (2001) respectively. CONCLUSIONS: Judicious application of eSET can halve the twinning rate while maintaining the overall pregnancy rate.     

Feeding NOD mice with pig splenocytes induces transferable mechanisms that modulate cellular and humoral xenogeneic reactions against pig spleen or islet cells

We have reported previously that oral administration of pig cells to NOD mice modified xenogeneic cellular response against pig islet cells (PICs), and hypothesized that it may have induced active suppression. This preliminary report evaluated only the effect of feeding pig cells by 'primary' proliferation, i.e. when splenocytes from fed mice are confronted with pig cells in vitro. The present study also considered 'secondary' proliferation and cytokine production after feeding and subsequent in vivo graft of pig cells. Additionally, serum IgM and IgG isotypes were quantified by ELISA using pig target cells. Induction of active mechanism by feeding was hypothetical, which led us here to transfer splenocytes from mice fed pig spleen cells (PSC) and evaluate 'primary' (after transfer) and 'secondary' (after transfer and subsequent graft of pig cells) proliferations and cytokine secretions in recipient mice. We also determined whether the effects of feeding pig cells persisted after depression of suppressor mechanisms by cyclophosphamide. Mice fed with PSC displayed increased 'primary' splenocyte proliferation to PSC or PIC (P < 0.0001), while 'secondary' responses were decreased (P < 0.03) in those fed PSC and subsequently grafted with PSC. The increased 'primary' and decreased 'secondary' proliferations were reduced (P < 0.04) by pretreatment with cyclophosphamide. The IL-10/ and IL-4/IFNgamma ratios produced in response to PSC increased (P < 0.04) in mice fed and grafted with PSC compared to those grafted only with PSC. IgM and IgG levels against pig cells were, respectively, increased (P < 0.04) and decreased (P < 0.04) in mice fed and grafted with PSC. IgG2a and IgG2b, but not IgG1, levels were lower (P < 0.01). These effects of feeding PSC on 'secondary' proliferation, cytokine and antibody productions, were not detected when mice were fed PSC only after graft with PSC. Transfer with splenocytes from mice fed PSC increased 'primary' proliferation of splenocytes from recipient mice in response to PSC (P < 0.02) or PIC (P < 0.05). After transfer with splenocytes from PSC-fed mice and graft with PSC, 'secondary' proliferation to pig cells were reduced (P < 0.04), and the IL-10/IFNgamma ratio produced in response to PSC was increased fourfold. Thus, oral administration of PSC induces active transferable mechanisms, characterized by a biphasic pattern with early increased 'primary' xenogeneic cellular reactions to both PSC and PIC, followed by decreased 'secondary' responsiveness and a concomitant shift of the Th1/Th2 balance towards greater Th2 influence. Decreased responsiveness may be due to active suppression, even though induction of anergy or deletion cannot be excluded.     

Serial cloning of pigs by somatic cell nuclear transfer: restoration of phenotypic normality during serial cloning.

Somatic cell nuclear transfer (scNT) is a useful way to create cloned animals. However, scNT clones exhibit high levels of phenotypic instability. This instability may be due to epigenetic reprogramming and/or genomic damage in the donor cells. To test this, we produced transgenic pig fibroblasts harboring the truncated human thrombopoietin (hTPO) gene and used them as donor cells in scNT to produce first-generation (G1) cloned piglets. In this study, 2,818 scNT embryos were transferred to 11 recipients and five G1 piglets were obtained. Among them, a clone had a dimorphic facial appearance with severe hypertelorism and a broad prominent nasal bridge. The other clones looked normal. Second-generation (G2) scNT piglets were then produced using ear cells from a G1 piglet that had an abnormal nose phenotype. We reasoned that, if the phenotypic abnormality of the G1 clone was not present in the G2 and third-generation (G3) clones, or was absent in the G2 clones but reappeared in the G3 clones, the phenotypic instability of the G1 clone could be attributed to faulty epigenetic reprogramming rather than to inherent/accidental genomic damage to the donor cells. Blastocyst rates, cell numbers in blastocyst, pregnancy rates, term placenta weight and ponderal index, and birth weight between G1 and G2 clones did not differ, but were significantly (P < 0.05) lower than control age- and sex-matched piglets. Next, we analyzed global methylation changes during development of the preimplantation embryos reconstructed by donor cells used for the production of G1 and G2 clones and could not find any significant differences in the methylation patterns between G1 and G2 clones. Indeed, we failed to detect the phenotypic abnormality in the G2 and G3 clones. Thus, the phenotypic abnormality of the G1 clone is likely to be due to epigenetic dysregulation. Additional observations then suggested that expression of the hTPO gene in the transgenic clones did not appear to be the cause of the phenotypic abnormality in the G1 clones and that the abnormality was acquired by only a few of the G1 clone's cells during its gestational development.