Characterization of somatic cell hybrids exhibiting extinction of AFP,Albumin and an AFP-HPRT Transgene

We utilized an AFP-HPRT transgene, i.e. the HPRT coding sequences under the regulation of AFP enhancer and promoter sequences, to localize the AFP extinguisher locus in intertypic somatic cell hybrids (hepatoma X fibroblast). This hybrid gene construct, which directly links AFP regulation to a reversibly selective gene, enabled the selection of stably transfected cells which express AFP, as well as cells showing extinction of AFP. Mouse hepatoma cells stably transfected with and expressing the transgene were fused to human fibroblasts, and the resulting somatic cell hybrids were characterized using Southern, Northern and karyotypic analyses. That several hybrids exhibited the proper extinction of AFP, AFP-HPRT and albumin suggests coregulation of these genes by an extinguisher. Segregant lines derived from these hybrids were selected for the loss of extinguisher activity and for reexpression of the transgene. Karyotypic analysis of hybrid and segregant lines, exhibiting proper AFP, albumin and AFP-HPRT phenotypes, revealed that the presence of human chromosome 7 was most closely associated with the AFP-extinguished state. The hybrids generated in these studies now make it possible to isolate the sequences responsible for AFP and albumin extinction.     

A case of the right subclavian artery as the last branch of the aortic arch in the human fetus and a new classification on these variations

An anomalous case of the right subclavian artery arising from the aortic arch as the last branch, in which the first branch was the right common carotid, the second the left common carotid and the third the left subclavian artery, was found in a 10 months human fetus among 173 fetuses. The right subclavian artery arose from the posterior wall of the aortic arch at the level of the Th4 and passed obliquely between the esophagus and the thoracic vertebrae. The right and the left vertebral arteries arising from the subclavian arteries on the same side entered the transverse foramen of the C6 of each side. This case belonged to type G of Adachi's classification and as well type 5 of Holzapfel's. The present authors wish to offer a new trial classification on these variations, including the origins and numbers of the vertebral arteries, by investigating many original reports in Japanese, as follows: 1) A new classification is fixed on the basis of the type G and H of Adachi-Williams et al.-Nakagawa in the classification of the branching types of the aortic arch. The type G represents that the right common carotid, the left common carotid, the left subclavian and the right subclavian arteries arise from the aortic arch in this order. The type H represents that the bicarotid trunk, the left subclavian and the right subclavian arteries arise from the aortic arch. 2) When the left vertebral artery arising from the aortic arch is found in the type G and H, "C" is prefixed G or H, as type CG, type CH. 3) When the right vertebral artery arising from the right common carotid artery is found, a prime mark, "', is put on G or H, as type G', type H'. 4) In order to represent a compound type of the above 2) and 3), both "C" and "' are put, as type CG', type CH'. 5) When the bilateral vertebral arteries arising from the respective subclavian artery are found in the above 2), 3) and 4) "2" postfixed "C" and the prime mark "', as type G'2, type C2G, type CG'2, type C2G', type C2G'2, type H'2, type C2H, type CH'2, type C2H', type C2H'2. According to the above new classification, Adachi's type G can be arranged into 18 branching types. This classification may be helpful and sufficient to provide more than 100 cases of the type G and H reported on Japanese.(ABSTRACT TRUNCATED AT 250 WORDS)     

L1 knockout mice show dilated ventricles, vermis hypoplasia and impaired exploration patterns

L1 is a neural cell adhesion molecule mainly involved in axon guidance and neuronal migration during brain development. Mutations in the human L1 gene give rise to a complex clinical picture, with mental retardation, neurologic abnormalities and a variable degree of hydrocephalus. Recently, a transgenic mouse model with a targeted null mutation in the L1 gene was generated. These knockout (KO) mice show hypoplasia of the corticospinal tract. Here we have performed further studies of these KO mice including magnetic resonance imaging of the brain, neuropathological analysis and behavioral testing. The ventricular system was shown to be abnormal with dilatation of the lateral ventricles and the 4th ventricle, and an altered shape of the Sylvius aqueduct. Additionally, the cerebellar vermis of the KO mice is hypoplastic. Their exploratory behavior is characterized by stereotype peripheral circling reminiscent of that of rodents with induced cerebellar lesions.      

Rapid detection of beta-globin gene (HBB) mutations coupling heteroduplex and primer-extension analysis by DHPLC

Beta-thalassemia is a common inherited disease, resulting from one or more of a total of more than 200 different mutations in the beta-globin gene (HBB). Efficient and reliable mutation-screening methods are essential in order to establish appropriate prevention programs for at-risk populations based upon a molecular diagnosis. We have developed a rapid and highly-specific mutation screening test for the diagnosis of beta-thalassemia by coupling heteroduplex and primer-extension analysis based on the denaturing high performance liquid chromatography (DHPLC) system. A total of 161 healthy heterozygous Taiwanese carriers featuring 10 different HBB mutations and 30 patients exhibiting 12 different compound heterozygous or homozygous HBB mutations were subjected to DHPLC. The elution profile for the heteroduplex analysis of DHPLC could be successfully used to identify the common disease-causing mutations of HBB. To further confirm the sequence variants, we developed a technique combining multiplex primer-extension analysis coupled with DHPLC for the genotyping of eight common disease-causing mutations in the HBB gene. Overall, by coupling heteroduplex and primer-extension analysis based upon DHPLC, we were able to unambiguously identify the most-common beta-thalassemia mutations corresponding to more than 99% of HBB alleles among the Taiwanese population. In conclusion, compared to classic approaches to mutation screening for this malady, we suggest that DHPLC is an excellent technique to be applied to the genetic screening of prenatal and postnatal individuals as a part of a diagnosis program for beta-thalassemia and provides a more-efficient, economic, and sensitive means to undertake such a screening program.     

Lupus-related advanced liver involvement as the initial presentation of systemic lupus erythematosus.

BACKGROUND AND PURPOSE: Systemic lupus erythematosus (SLE), a prototype of systemic autoimmune disease characterized by multiorgan involvement with diverse clinical and serological manifestations, principally affects women in their child-bearing years. Clinically significant hepatic abnormality as the initial presentation of SLE has rarely been reported. METHODS: Eleven patients with lupus with initial presentation of lupus-related hepatitis were included in this retrospective review. Clinical manifestation, immunological profiles, and risk factors for poor prognosis were analyzed. RESULTS: The most commonly associated clinical manifestations were found to be thrombocytopenia, leukopenia, advancing age, and presence of anti-SSA/Ro antibody and anti-thyroid antibodies. The diagnosis of SLE was delayed due to dominant hepatic abnormalities. Age greater than 50 years and marked hepatic decompensation in accordance with Child classification B and C might suggest poor prognosis (p=0.06). However, the p value was not statistically significant because of the small sample size. CONCLUSIONS: Lupus-related hepatitis, particularly in late-onset lupus, is common. In addition, the presence of anti-SSA, anti-thyroglobulin, and anti-microsomal antibodies is indicative of hepatic involvement in patients with SLE.     

Decreased production of CD8 (T8) antigen after immunotherapy

The working mechanism(s) of immunotherapy still remains ill defined. As T cells bearing CD8 antigen possess suppressor/cytotoxic function, this study was conducted to examine the effect of immunotherapy on the production of CD8 antigen. Peripheral blood mononuclear cells (MNC) were obtained from 21 newly diagnosed and 23 hyposensitized (>1 year) asthmatic children and 13 agematched normal children. MNC were stimulated with crude mite extract (Dermatophagoides farinae) for 7 days and with phytohemagglutinin and concanavalin A for 3 days. The CD8 antigen and interleukin-2 receptor (IL-2R) in plasmas and culture supernatants were measured by CELLFREE T8 and IL-2R test kits (T Cell Sciences, USA). The results showed the following. (1) Plasma CD8 antigen was markedly increased in new patients compared to normals (536.7 ± 212.3 vs 222.5 ± 104.0 units/ml;P<0.001) and decreased to normal after immunotherapy (275.7 ± 98.5 units/ml). (2) When stimulated with mite allergen, MNC from both new and hyposensitized patients produced a much greater amount of CD8 antigen compared to those from normals. However, after immunotherapy MNC tended to produce less CD8 antigen, although not to a significant degree. (3) No difference in CD8 antigen production was seen among three groups when lymphocytes were stimulated with mitogens. (4) Production of CD8 antigen paralleled that of IL-2R. Thus, CD8 production was specifically decreased after immunotherapy and this fact reflects a hyposensitized state of T cells after long-term, repeated injection of allergens.     

四川参叶的生药学鉴定研究

参叶为少常用中药,主产于四川。我们通过调查,发現四川药用的参叶共有8种,均属人参属植物。本文报道了对8种参叶的生药学鉴定研究结果,提出了它们之间性状、显微特征的异同,创设了表皮细胞数的概念,并肯定了它在生药鉴定中的价值。研究结果对于合理用药、开发利用人参属植物资源及人参属植物的分类均有一定的价值。     

Two-stage Study of Familial Prostate Cancer by Whole-exome Sequencing and Custom Capture Identifies 10 Novel Genes Associated with the Risk of Prostate Cancer

BackgroundFamily history of prostate cancer (PCa) is a well-known risk factor, and both common and rare genetic variants are associated with the disease.ObjectiveTo detect new genetic variants associated with PCa, capitalizing on the role of family history and more aggressive PCa.Design, setting, and participantsA two-stage design was used. In stage one, whole-exome sequencing was used to identify potential risk alleles among affected men with a strong family history of disease or with more aggressive disease (491 cases and 429 controls). Aggressive disease was based on a sum of scores for Gleason score, node status, metastasis, tumor stage, prostate-specific antigen at diagnosis, systemic recurrence, and time to PCa death. Genes identified in stage one were screened in stage two using a custom-capture design in an independent set of 2917 cases and 1899 controls.Outcome measurements and statistical analysisFrequencies of genetic variants (singly or jointly in a gene) were compared between cases and controls.Results and limitationsEleven genes previously reported to be associated with PCa were detected (ATM, BRCA2, HOXB13, FAM111A, EMSY, HNF1B, KLK3, MSMB, PCAT1, PRSS3, and TERT), as well as an additional 10 novel genes (PABPC1, QK1, FAM114A1, MUC6, MYCBP2, RAPGEF4, RNASEH2B, ULK4, XPO7, and THAP3). Of these 10 novel genes, all but PABPC1 and ULK4 were primarily associated with the risk of aggressive PCa.ConclusionsOur approach demonstrates the advantage of gene sequencing in the search for genetic variants associated with PCa and the benefits of sampling patients with a strong family history of disease or an aggressive form of disease.Patient summaryMultiple genes are associated with prostate cancer (PCa) among men with a strong family history of this disease or among men with an aggressive form of PCa.