2,8-Dihydroxyadenine urolithiasis in a patient with considerable residual adenine phosphoribosyltransferase activity in cell extracts but with mutations in both copies of APRT

We have examined the mutational basis of adenine phosphoribosyltransferase (APRT, EC 2.4.2.7) deficiency (MIM 102600) in a patient of Polish origin who has been passing 2,8-dihydroxyadenine (DHA) stones since birth, but has considerable residual enzyme activity in lymphocyte extracts. The five exons and flanking regions of APRT were amplified by PCR and then sequenced. A single T insertion was identified at the intron 4 splice donor site (TGgtaa to TGgttaa:IVS4+2insT) in one allele from the proband, his mother, and brother. A G-to-T transversion in exon 5 (GTC-to-TTC:c.448G>T, V150F) was identified in the other allele, and this mutation was also present in one allele from the father and the paternal grandmother. Tru91 and AvaII digestions of PCR products spanning exons 4 and 5, respectively, confirmed the mutations. The mother was heterozygous for an intragenic TaqI site, but all other family members were homozygous for the presence of this site. IVS4+2insT, located on the allele containing the TaqI site, has been identified previously in several families from Europe, suggesting a founder effect, but the substitution in exon 5 is a novel mutation. IVS4+2insT is known to result in complete loss of enzyme activity, and our results suggest that V150F produces an enzyme that is nonfunctional in vivo but has considerable residual activity in vitro.     

Derivation of electron and photon energy spectra from electron beam central axis depth dose curves

A method for deriving the electron and photon energy spectra from electron beam central axis percentage depth dose (PDD) curves has been investigated. The PDD curves of 6, 12 and 20 MeV electron beams obtained from the Monte Carlo full phase space simulations of the Varian linear accelerator treatment head have been used to test the method. We have employed a 'random creep' algorithm to determine the energy spectra of electrons and photons in a clinical electron beam. The fitted electron and photon energy spectra have been compared with the corresponding spectra obtained from the Monte Carlo full phase space simulations. Our fitted energy spectra are in good agreement with the Monte Carlo simulated spectra in terms of peak location, peak width, amplitude and smoothness of the spectrum. In addition, the derived depth dose curves of head-generated photons agree well in both shape and amplitude with those calculated using the full phase space data. The central axis depth dose curves and dose profiles at various depths have been compared using an automated electron beam commissioning procedure. The comparison has demonstrated that our method is capable of deriving the energy spectra for the Varian accelerator electron beams investigated. We have implemented this method in the electron beam commissioning procedure for Monte Carlo electron beam dose calculations.     

白细胞介素1,肿瘤坏死因子α和脂多糖对人脐静脉内皮细胞表达 …

目的 观察细胞因子白细胞介素１（ＩＬ－１）β、肿瘤坏死因子（ＴＮＦ）α和脂多糖（ＬＰＳ）是否诱导人脐静脉内皮细胞表达单核细胞趋化蛋白１（ＭＣＰ－１）ｍＲＮＡ及蛋白。方法 选取生长汇合的人脐胸脉内皮细胞,在其培养基中分别加入终浓度为２ｎｇ／ｍｌ的ＩＬ－１β、２０ｎｇ／ｍｌ的ＴＮＦβ和１００ｎｇ／ｍｌ的ＬＰＳ,３７℃共育４ｈ后,按照一步法提取其总ＲＮＡ,用γ－＾２２Ｐ标记的寡核苷酸ｄｏｔ ｂｌｏｔ分析     

A model of corrective gene transfer in X-linked ichthyosis

Single gene recessive genetic skin disorders offer attractive prototypes for the development of therapeutic cutaneous gene delivery. We have utilized X-linked ichthyosis (XLI), characterized by loss of function of the steroid sulfatase arylsulfatase C (STS), to develop a model of corrective gene delivery to human skin in vivo. A new retroviral expression vector was produced and utilized to effect STS gene transfer to primary keratinocytes from XLI patients. Transduction was associated with restoration of full-length STS protein expression as well as steroid sulfatase enzymatic activity in proportion to the number of proviral integrations in XLI cells. Transduced and uncorrected XLI keratinocytes, along with normal controls, were then grafted onto immunodeficient mice to regenerate full thickness human epidermis. Unmodified XLI keratinocytes regenerated a hyperkeratotic epidermis lacking STS expression with defective skin barrier function, effectively recapitulating the human disease in vivo. Transduced XLI keratinocytes from the same patients, however, regenerated epidermis histologically indistinguishable from that formed by keratinocytes from patients with normal skin. Transduced XLI epidermis demonstrated STS expression in vivo by immunostaining as well as a normalization of histologic appearance at 5 weeks post-grafting. In addition, transduced XLI epidermis demonstrated a return of barrier function parameters to normal. These findings demonstrate corrective gene delivery in human XLI patient skin tissue at both molecular and functional levels and provide a model of human cutaneous gene therapy.      

γ-氨基丁酸受体拮抗剂与海马齿状回颗粒细胞核转录因子κB的激活

目的:研究γ-氨基丁酸(GABA)受体拮抗剂荷包牡丹碱及木防己苦毒素对核转录因子κB(NF-κB)的激活和调控作用。方法:利用海马脑片培养技术和免疫荧光技术,对经GABA受体拮抗剂处理的NF-κB激活细胞进行了计数和统计学检验。结果:经GABA受体拮抗剂处理后,海马齿状回颗粒细胞NF-κB细胞激活率高于对照组。结论:GABA受体拮抗剂可能通过GABA受体参与对NF-κB信号转录途径激活和调控,最终影响某些基因转录和表达。     

RNAi抑制XJ-160病毒复制的研究

目的通过建立RNA干扰(RNA interference，RNAi)抑制XJ-160病毒复制的模型，在序列特异性、位置效应和量效关系等方面，探讨RNAi对XJ-160病毒复制的影响，为进一步研究RNAi的抗病毒作用和机制奠定基础。方法按照siRNA(short interferencing RNA)的设计要求合成XJ-160病毒特异siRNA，脂质体法转染BHK-21细胞后感染XJ-160病毒，通过测定病毒滴度、蛋白表达量及XJ-160病毒：RNA合成研究siRNA对XJ-160病毒复制的抑制。结果成功建立了RNAi抑制XJ-160病毒复制的模型，探讨了RNAi抑制该病毒复制作用的特点。结论外源导入的siRNA能够抑制XJ-160病毒的复制，并且这种抑制作用具有明显的序列特异性、位置效应和存在量效关系等特点；siRNA是通过降解病毒RNA实现抑制病毒复制作用的。     

Production and Characterization of Generic Antibodies Against s-Triazine and Sulfonylurea Herbicides

Two different hapten designs have been suggested for production of generic antibodies. The first approach was based on the immunogen prepared by conjugating simazine mimic to carrier proteins through the Cl position of the s-triazine herbicide. For sulfonylurea herbicides a single ring hapten strategy was designed in order to produce antibodies with dominant selectivity towards arylsulfonyl or triazine moieties of metsulfuron-methyl. The best monoclonal antibody raised against simazine-derived hapten mimic (clone B10/B8/D2) exhibited in direct ELISA a broad pattern for a group of methylthio and methoxy s-triazines. Cross-reactivities based on simazine ( = 100%) were 417, 125, 25, 50, 28, 42 and 11% for simetryn, ametryn, prometryn, terbutryn, aziprotryn, atraton and atrazine, respectively. The superiour generic pattern was found for monoclonal antibody raised against s-triazine moiety of metsulfuron-methyl herbicide (clone 2C8/C8). This antibody showed in indirect ELISA the cross-reactivity values 100, 142, 95, 60, 40, 167, 83 and 21% for metsulfuronmethyl, cinosulfuron, triasulfuron, primisulfuron-methyl, thifensulfuron-methyl, chlorsulfuron and prosulfuron and tribenuron-methyl, respectively. The assays using both monoclonal and polyclonal antibodies operated within the ppt-ppb calibration ranges.     

鹅胃肠内分泌细胞的免疫组织化学定位

应用６种胃肠激素抗血清，对鹅胃肠各段的内分泌细胞分布进行了免疫组织化学定位。腺胃显示有较多促胃泌素释放肽和生长抑素细胞。肌胃有较多促胃泌素释放肽、生长抑素和胃泌素细胞，少量胰多肽细胞。幽门部有大量的生长抑素细胞，密集的胃泌素细胞，偶见胰多肽细胞。小肠内有胃泌素、胰多肽和生长抑素细胞，细胞类型和数量由前段向后段逐渐减少。未检出胃动素和抑胃肽细胞。     

Parental origin and mechanism of formation of X chromosome structural abnormalities: Four cases determined with RFLPs

Parental origin and mechanism of formation of X chromosome structural anbormalities were studied in one each case of dup(X)(pter p11.4::p22.1qter), del(X)(qterp11:), i(X)(qtercenqter), and inv dup(X) (pterq22::q22pter) using various X-linked RFLPs as genetic markers. Segregation and densitometric analyses on polymorphic DNAs revealed that the dup(Xp) and the del(Xp) are both of paternal origin and the i(Xq) and i dic(X) are of maternal origin. The dup(Xp) had arisen by an unequal sister chromatid exchange and the del(Xp) had occurred through an intrachromosomal breakage-reunion mechanism, both in the paternal X chromosome. The i(Xq) had arisen either through centromere fission of a maternal X chromosome, followed by duplication, of its long-arm, or through a translocation between two maternal X chromosomes after meiotic crossing-over. The inv dup(X) arose through sister chromatid breakage and reunion in a maternal X chromosome. These results, together with those of previous studies, suggest that thede novo abnormalities due to events involving centromere disruption arise predominantly during oogenesis, while those due to simple breakage-reunion events occur preferentially during spermatogenesis.