Genotype-phenotype correlation and frequency of the 3199del6 cystic fibrosis mutation among I148T carriers: results from a collaborative study.

PURPOSE: We expect that the mutation panel currently recommended for preconception/prenatal CF carrier screening will be modified as new information is learned regarding the phenotype associated with specific mutations and allele frequencies in various populations. One such example is the I148T mutation, originally described as a severe CF mutation. After implementation of CF population-based carrier screening, we learned that I148T exists as a complex allele with 3199del6 in patients with clinical CF, whereas asymptomatic compound heterozygotes for I148T and a second severe CF mutation were negative for 3199del6. METHODS: We performed reflex testing for 3199del6 on 663 unrelated specimens, including I148T heterozygotes, compound heterozygotes, and a homozygous individual. RESULTS: Less than 1% of I148T carriers were also positive for 3199del6. Excluding subjects tested because of a suspected or known CF diagnosis or positive family history, 0.6% of I148T-positive individuals were also positive for 3199del6. We identified 1 I148T homozygote and 6 unrelated compound heterozygous individuals with I148T and a second CF variant (2 of whom also carried 3199del6). In addition, one fetus with echogenic bowel and one infertile male were heterozygous for I148T (3199del6 negative). CONCLUSIONS: Reflex testing for 3199del6 should be considered whenever I148T is identified. Reflex testing is of particular importance for any symptomatic patient or whenever one member of a couple carries a deleterious CF mutation and the other member is an I148T heterozygote. Further population data are required to determine if I148T, in the absence of 3199del6, is associated with mild or atypical CF or male infertility.     

Expression and functional analysis of Tgif during mouse midline development.

The Tgif gene encodes a homeodomain protein that functions as a transforming growth factor beta (TGF-beta) repressor by binding to Smad2. Mutations in the TGIF gene are associated with human holoprosencephaly, a common birth defect caused by the failure of anterior ventral midline formation. However, Smad2-mediated TGF-beta signaling in the axial mesendoderm has been demonstrated to be essential for ventral midline formation, and loss of a Smad2 antagonist should in principle promote rather than inhibit ventral midline formation. This suggests a more complex mechanism for the function of TGIF in controlling ventral midline formation. To explore the role of TGIF in ventral forebrain formation and patterning, we investigated Tgif expression and function during mouse development by in situ hybridization and gene targeting. We found that Tgif is highly expressed in the anterior neural plate, consistent with the proposed neural differentiation model in which TGF-beta suppression is required for normal neural differentiation. This result suggests a possible role for Tgif in anterior neural differentiation and patterning. However, targeted disruption of the Tgif gene during mouse development does not cause any detectable defects in development and growth. Both histological examination and gene expression analysis showed that Tgif-/- embryos have a normal ventral specification in the central nervous system, including the forebrain region. One interpretation of these results is that the loss of TGIF function is compensated by other TGF-beta antagonists such as c-Ski and SnoN during vertebrate anterior neural development.     

Expression of mucin genes in the human testis and its relationship to spermatogenesis

In this study we investigate the expression pattern of mucin genes in the human testis and evaluate the relationship between the expression of mucin genes and impaired spermatogenesis in the human testis. Thirty human testis tissues were collected from patients undergoing diagnostic testicular biopsy to investigate the cause of infertility. One part of the tissue underwent histological observation, and the other part of the tissue was subjected to semiquantitative RT-PCR of mucin genes, that is, mucin1, 2, 3, 4, and 9. The relative amount of mucin mRNAs was calculated by densitometry using glyceraldehydes-3-phosphate dehydrogenase (GAPDH) as an internal control. The samples were histologically diagnosed as either obstructive azoospermia with normal spermatogenesis (n = 13) or non-obstructive azoospermia with impaired spermatogenesis (n = 17). In the human testis with normal spermatogenesis, mRNA expression of mucin1, 9, 13 and GAPDH were found, but RT-PCR products of mucin 2, 3 and 4 were not detected. In the testis with impaired spermatogenesis, however, RT-PCR product of mucin1 was not found. There was no difference in the other mucin mRNA expression patterns between the testis with either normal or impaired spermatogenesis. To our knowledge, this study is the first that has detected the mRNA of mucin9 and 13 in human testis. This study also shows that mucin1 expression might be closely related to spermatogenesis. Our findings should be substantiated by more direct evidence, such as mucin protein expression and localization.     

Apoptosis of malignant cells in Hodgkin's disease is related to expression of the cdk inhibitor p27KIP1

Previous results from B-cell chronic lymphocytic leukaemia suggest that expression of p27KIP1 might be important in protection from apoptosis. Given the relevance of apoptosis to the pathogenesis of Hodgkin's disease (HD), it was decided to examine the expression of p27KIP1 in relation to apoptosis in these lesions. Paraffin-wax sections from a total of 65 histologically confirmed HD tumours were used to derive apoptotic index (AI) and DNA fragmentation index (DFI) scores, which were compared with the expression of various cell-cycle-regulating proteins, including p27KIP1 (p27), p21WAF1/CIP1 (p21) and cyclin D1, and with Epstein-Barr virus (EBV) status. The DFI was measured by TdT-mediated dUTP-FITC nick end-labelling (TUNEL), and the AI by conventional morphology. Cells showing the typical morphology of apoptosis, together with those resembling so-called 'mummified' Hodgkin/Reed-Sternberg (HRS) cells, were included in AI measurements. Increasing numbers of p27-positive HRS cells were associated with lower levels of apoptosis in these cells, as indicated by significantly lower AI and DFI scores. There was a trend towards poorer survival in those patients with the highest numbers of p27-positive HRS cells and with lower AI and DFI scores, but these differences were not statistically significant. p21-positive HRS cells were significantly more frequent in those cases with lower AI scores. A similar trend was observed for p21 and DFI, although this relationship was not statistically significant. There was also a trend towards higher levels of cyclin D1 protein in HD cases with high AI and DFI values. A tendency for increasing numbers of p27-positive and p21-positive HRS cells in EBV-positive cases was noted, but this relationship was not statistically significant. EBV status did not correlate with either AI or DFI scores. The results of this study suggest that p27, and possibly also p21, may be involved in protection from apoptosis in HD.     

A case of Klinefelter syndrome with retroperitoneal teratoma

Klinefelter syndrome (KS) is often associated with various neoplasms, especially germ cell tumors. Mediastinum is the most favored site of extragonadal germ cell tumors with KS, which is somewhat different from those without KS. The retroperitoneal germ cell tumor in KS is very rare. A five-month-old boy with an abdominal mass was found to have a retroperitoneal tumor. After surgical removal, he was diagnosed to have mature cystic teratoma. Cytogenetic study of his peripheral lymphocytes revealed that his karyotype was consistent with KS. This case suggests that patients with KS might be at risk of having germ cell tumors in sites other than mediastinum. It also suggests that all cases with these tumors should be screened for the presence of karyotypic abnormalities, and it might help to assess the exact correlation between germ cell tumors and KS, and to treat them accordingly.     

Clinical and cytogenetics studies on acute myeloid leukemia with abnormality of chromosome 11

OBJECTIVE: To investigate the incidence of chromosome 11 abnormality in acute myeloid leukemia and its relationship with the clinical aspects and prognosis. METHODS:Conventional cytogenetic analysis of R-band was used to detect the abnormalities of chromosome 11 in 356 acute myeloid leukemia patients. RESULTS: Thirty-four out of 356 patients (9.55%) had abnormalities of chromosome 11, of which 20 (58.8%) involved in 11q23, 7 (19.9%) had translocations involving 11p15, 5 (14.7%) had-11, and the rest had other abnormalities such as +11, and t(11;14). The incidence of 11q23 involvement in M4 and M5 was higher than other subtypes of acute myeloid leukemia (AML). Ten cases with 11q23 abnormality had additional cytogenetic aberrations. In 30 cases treated with chemotherapy, 13 cases acquired complete remission (CR). The CR rate was lower than that of whole cases of acute myeloid leukemia(34.3% versus 64.0%). The CR rate of AML with 11q23 abnormality was lower than that of AML with normal karyotype (25% versus 55.6%). In other 10 patients with additional chromosome aberrations, the CR rate was lower than that of AML with 11q23 alone. In 7 patients with translocations at 11p15, only 3 patients acquired CR, and 2 patients relapsed early. Only 2 patients acquired CR in 5 patients with-11. CONCLUSION: 11q23 was a frequent aberration in chromosome 11 anomaly, which was often detected in M4 and M5. It might be associated with the pathogenesis of acute monolytic leukemia. The patients with chromosome 11 anomaly had poorer prognosis.     

Genetic control of the antibody response to poly(L Tyr, L Glu)-poly(DL Ala)--poly(L Lys) in mice: analysis of (low responder x low responder)F1 hybrids

The antibody response to the synthetic polypeptide poly (L Tyr, L Glu)-poly (DL Ala)--poly (L Lys) designated (T,G)-A--L, was investigated in inbred, congenic, F1 and F2 hybrid strains of mice. The antibody response was analysed at both low (10 microgram) and high (50 microgram) immunizing doses of (T,G)-A--L. Antibodies were measured using both a modified Farr assay and a plate binding assay. At low immunizing doses it was found that all of the congenic and non-congenic (low responder x low responder) F1 hybrids were low responders. However, the quantitative antibody response of one non-congenic (low responder x low responder) F2 hybrid segregated in a 1:1 ratio of high responders to low responders, suggesting some form of complementation of (T,G)-A--L Ir genes. At high immunizing doses it was found that congenic and non-congenic (low responder x low responder) F1 hybrids were all high responders, indicating a complementation of Ir genes to (T,G)-A--L. This complementation was confirmed using two different routes of immunization, namely footpad and intraperitoneal. Furthermore the quantitative antibody responses of (low responder x low responder) F2 hybrids segregate in a 1:1 ratio of high responders to low responders. The class of antibodies produced to (T,G)-A--L in (low responder x low responder) F1 hybrids was determined by gel filtration on Sephadex G-200, and found to be predominantly IgG, with lesser amounts of IgM.     

Simulating activations with cytoarchitecture

Cytoarchitectonic delineation of areas in post-mortem human brains provides the precise location of these areas. It has been possible to study the size and location of areas between post-mortem brains with multi-subject cytoarchitectonic data. If the structure–function relationship is assumed to be a one-to-one mapping for the purposes of inter-subject variability, then functional areas in the cortex will also adhere to the structure, and therefore, the location and size of cytoarchitectonic areas in the brain. Thus, it is possible to use the cytoarchitectonic data as being representative of the size and location of functional activations. Under this assumption, we simulated activations in cytoarchitectonic areas from ten post-mortem brains in this study. We then treated these data as we would a normal PET experiment. The purpose of this study is to demonstrate a standard PET image analysis on a simulated ten-subject PET study using cytoarchitecture to localize the activations. By doing so, we simulate activations with real inter-subject variability with the size and location of each area. Significant activations were obtained for activations simulated in areas 3a and 3b. A voxel-wise conjunction between simulated data and experimental data was made to better determine the underlying areas activated by the experimental tasks. This study presents a novel technique for demonstrating the effect of standard image analysis on the location and size of simulated activations as determined by cytoarchitectonic data from multiple subjects. Furthermore, this technique has been applied to better determine the underlying areas activated in an experiment.     

无精子症及严重少精子症患者Y染色体AZF微缺失及点突变的研究

目的通过对无精子症和严重少精子症患者Y染色体AZF微缺失的检测,确定AZF微缺失的发生率及高发位点;同时,对正常生育男性、无精子症及严重少精子症患者进行sY254、sY255点突变检测,从分子水平探讨精子发生的机制,建立基因型与表型的关系。方法多重PCR技术对Y染色体上AZF 4个区域内的15个序列标签位点进行微缺失检测,采用SSCP方法进行sY254、sY255点突变检测;结果无精子症和严重少精子症患者Y染色体AZF微缺失率分别为9.80%和9.68%,显著高于少精子症组的3.34%(P<0.05);其中sY152、sY239、sY243、sY254、sY255在Y染色体上的位置相互毗邻,其联合缺失39例,占总缺失率的65.0%,AZFb+AZFd+AZFc联合缺失的有8人,占总缺失的13.3%;本研究中,正常生育男性、无精子症及严重少精子症患者均未发现sY254、sY255的点突变。结论Y染色体AZF微缺失是导致无精子症和严重少精子症的重要因素,sY152、sY239、sY243、sY254、sY255联合缺失是AZF的缺失热点;未发现sY254、sY255点突变。     

距骨颈骨折疗效分析

目的探讨距骨颈骨折治疗方案的选择以及疗效相关因素分析。方法距骨颈骨折15例，男13例，女2例。年龄16～63岁，平均40．5岁，合并腰椎、髋臼、跟骨、外踝骨折各1例。开放骨折2例。陈旧性骨折2例（〉1个月）。按Hawkins骨折分类：Ⅰ型3例、Ⅱ型9例、Ⅲ型3例。Ⅰ型骨折采用膝下管型石膏固定，其中1例复诊中因移位行切开复位松质骨螺钉固定。Ⅱ型骨折均行切开复位，克氏针或松质骨螺钉固定。Ⅲ型骨折1例开放性骨折，急诊行清创缝合，手法＋撬拔复位，脱位整复。一周后在C臂机引导下，采用二枚直径4．0mm中空螺钉由后向前固定距骨颈，1例急诊手术切开复位，克氏针内固定。结果本组13例获得随访，时间4～24个月，平均12.3个月。13例中发生距骨体缺血性坏死5例，其中Ⅱ型骨折4例，Ⅲ型骨折1例。另2例未统计在内。1例外院病例术后并发伤口感染长期不愈，行小腿截肢术。1例患者拒绝手术治疗。按Hawkims疗效标准，优4例，良4例，可3例，差2例，优良率69％。结论对无移位的距骨颈骨折，行膝下管型石膏固定6～12周，并定期拍片复查，距骨颈骨折脱位应急诊复位和可靠的内固定。以2枚拉力螺钉内固定为宜，尽量减少并发症。对距骨体缺血性坏死，或创伤性关节炎，则根据患者的实际情况采取相应治疗措施。