Prognostic significance of loss of heterozygosity on chromosome 9p21–22 in human esophageal cancer

Background Deletions involving chromosome 9p21, on which the tumor suppressor genep16/MTS1 is located, have been noted in esophageal cancer. We investigated the relationship between the deletion of chromosome 9p21–22 and the clinical features of esophageal cancer. Methods We examined the loss of heterozygosity (LOH) on chromosome 9p21–22 in 56 esophageal cancers using polymerase chain reaction (PCR) analysis and 2 microsatellite markers (RPS6 and IFNA). Results In 18 out of 50 informative cases (36%), LOH had occurred at 1 or 2 loci on chromosome 9p21–22. We found no relationship between LOH on chromosome 9p21–22 and patient sex, age tumor length, location, histologic differentiation, depth of tumor invasion, the extent of lymph node metastasis, histologic stage, or curability. Among 35 patients without an absolute noncurative resection, the mean survival of 11 patients with LOH on chromosome 9p21–22 was 19.3 months, compared with 42.3 months for 24 patients with a normal allele; thus, the survival rate of those with LOH was significantly lower than that of patients without LOH on chromosome 9p21–22 (log-rank test;P=0.03). Conclusion These data suggest that LOH on chromosome 9p21–22, on which the cell-cycle regulatorp16/MTS1 gene is located, may be related to cancer development, and probably can serve as a clinical marker for evaluating a patient's prognosis.     

可变数串联重复序列检测异基因外周血造血干细胞移植后植？…

目的 从血型、染色体核型、ＤＮＡ可变数串联重复序列（ＶＮＴＲ）几个方面进行对比研究以更好地评估异基因移植后的植入状态及其与疾病复发的关系。方法 采用聚合酶链反应（ＰＣＲ）方法扩增Ｄ１７Ｓ３０、Ｄ１Ｓ８０、ＡｐｏＢ３个不同位点的多态性,对１５例白血病患者异基因外周血造血干细胞移植（ａｌｌｏ－ＰＢＳＣＴ）后的嵌合状态进行检测,对其中７例异性间移植和３例血型不合移植又分别进行了染色体核型分析和全套血型分     

耳炎差异球菌与成人分泌性中耳炎的相关性研究

目的探讨耳炎差异球菌与成人分泌性中耳炎的关系。方法采用多重PCR的方法，检测39例(42耳)成年分泌性中耳炎患者的中耳积液中耳炎差异球菌以及三种常见细菌[肺炎链球菌(S．pneumoniae)、流感嗜血杆菌(H．influenzae)和卡他莫拉杆菌(M．catarrhalis)]的DNA，就细菌DNA存在与成人分泌性中耳炎患者上呼吸道感染(简称上感)史、病程、积液性质和抗生素服用史之间的关系进行分析。结果收集到的42份中耳积液标本中有5份(11．9％)检测到有耳炎差异球菌DNA存在。急性分泌性中耳炎者有1例(3．6％)测得耳炎差异球菌DNA存在，慢性者则有4例(28．6％)，两者差异有显著性意义。浆液性中耳积液有3例(13．0％)测得耳炎差异球菌DNA存在，粘液性中耳积液则有2例(10．5％)为阳性，两者差异有显著性意义。抗生素使用史对耳炎差异球菌DNA检出率的影响无统计学意义。结论细菌参与部分成人分泌性中耳炎的发病，耳炎差异球菌可能是其致病菌之一；且慢性分泌性中耳炎中耳积液的持续存在可能与耳炎差异球菌感染有关。     

实时定量PCR检测体外诱导条件下小鼠骨髓基质细胞S100mRNA表达水平变化

目的:检测S100 mRNA含量,初步探讨骨髓基质细胞体外诱导条件下向雪旺细胞样细胞分化的可能性。方法:采用差速贴壁的方法分离、培养小鼠骨髓基质细胞。经β-ME,ATRA,Forskolin,bFGF,PDGF,HRG体外诱导后,利用实时定量PCR检测S100 mRNA水平的变化。结果:诱导后,骨髓基质细胞S100 mRNA水平上升,诱导前后水平变化有统计学意义(P<0.05)。结论:实时定量PCR检测S100 mRNA含量具有较高的灵敏度和特异性。体外诱导后骨髓基质细胞S100 mRNA表达量增多。     

Rapid genotyping of newborn gene mutant mice

One important aspect of utilizing transgenic mice is the need to genotype them in order to distinguish mice that carry a disrupted gene or a transgene from mice that do not. Current methods for genotyping include isolation of genomic DNA from tail biopsies followed by PCR amplification. Particularly, both digestion of tail tissue using proteinase K as well as resuspension of purified DNA are time-consuming and were usually carried out overnight. Here, we describe a rapid and robust method for the genotyping of bdnf targeted mice which allows us to determine the genotype of newborn mice at the day of birth within 6 h. After a freezing–thawing step tail tissue is digested in less than 2 h, and the DNA is precipitated, resuspended and ready for PCR in about 60 min. The method could be easily adapted to a variety of different mutant mice and especially should benefit neuroscientists interested in using animals with known genotype very early in postnatal development.     

Molecular monitoring of tumour load kinetics predicts disease progression after non-myeloablative allogeneic stem cell transplantation in multiple myeloma.

BACKGROUND: Non-myeloablative allogeneic stem cell transplantation followed by immunomodulatory therapies is considered a potentially curative approach in the treatment of multiple myeloma and most effective in a minimal residual disease setting. PATIENTS AND METHODS: The aim of this study was to find the most sensitive real-time PCR assay (TaqMan), based on the IGH rearrangement, to quantify the tumour load of 11 patients with multiple myeloma after non-myeloablative allogeneic transplantation. Patient-allele specific primers (ASO) and the TaqMan probe were derived from CDR2 and CDR3 hypervariable regions of IGH, while consensus primers were located within the FR3 and FR4/JH regions. Four different approaches of primer combinations were tested. RESULTS: ASO-forward and -reverse primers together with the clone-specific TaqMan probe were the most sensitive approach compared with the JH (P=0.071) or the FR3 consensus primer (P <0.001). The detection limit amounted to 1/10(4)-1/10(5) cells. Consecutively, 120 samples from 11 patients prior and post allogeneic transplantation were analysed. Three patients reached complete clinical remission accompanied by molecular remission. Disease progression or relapse was seen in six patients. In five, molecular progressive disease was detected prior to the clinical diagnosis of progression or relapse. CONCLUSION: Patient-specific real-time IGH-PCR provides the opportunity for earlier treatment intervention.     

SARS-CoV-2 Variants in Paraguay: Detection and Surveillance with an Economical and Scalable Molecular Protocol

SARS-CoV-2 variant detection relies on resource-intensive whole-genome sequencing methods. We sought to develop a scalable protocol for variant detection and surveillance in Paraguay, pairing rRT-PCR for spike mutations with Nanopore sequencing. A total of 201 acute-phase nasopharyngeal samples were included. Samples were positive for the SARS-CoV-2 N2 target and tested with the Spike SNP assay to detect mutations associated with the following variants: alpha (501Y), beta/gamma (417variant/484K/501Y), delta (452R/478K), and lambda (452Q/490S). Spike SNP calls were confirmed using amplicon (Sanger) sequencing and whole-genome (Nanopore) sequencing on a subset of samples with confirmed variant lineages. Samples had a mean N2 Ct of 20.8 (SD 5.6); 198/201 samples (98.5%) tested positive in the Spike SNP assay. The most common genotype was 417variant/484K/501Y, detected in 102/198 samples (51.5%), which was consistent with the P.1 lineage (gamma variant) in Paraguay. No mutations (K417 only) were found in 64/198 (32.3%), and K417/484K was identified in 22/198 (11.1%), consistent with P.2 (zeta). Seven samples (3.5%) tested positive for 452R without 478K, and one sample with genotype K417/501Y was confirmed as B.1.1.7 (alpha). The results were confirmed using Sanger sequencing in 181/181 samples, and variant calls were consistent with Nanopore sequencing in 29/29 samples. The Spike SNP assay could improve population-level surveillance for mutations associated with SARS-CoV-2 variants and inform the judicious use of sequencing resources.     

套式聚合酶链反应及限制酶分析检测新生儿肝炎巨细胞病毒感染

在人巨细胞病毒（HCMV）AD169株直接早期蛋白EcoRIJDNA片段内，自行设计二对引物，建立套式PCR检测HCMVDNA。经实验证明有高度的特异性与敏感性，对其它疙疹类病毒和正常人基因组DNA无交叉反应。一次性PCR检测HCMVAD169株基因组的最小检出量为100fg，而套式PCR可达10fg，经PStI酶切后，得到的片段与设计相符。对23例新生儿肝炎临床标本进行病毒分离，一次性PCR，套式PCR检测，结果表明套式PCR能进一步提高一次性PCR的特异性与敏感性，适用于新生儿肝炎HCMV感染的临测。     

Sleep-promoting activity of lotus (Nelumbo nucifera) rhizome water extract via GABAA receptors

ContextThe sleep-promoting activity of Nelumbo nucifera Gaertn. (Nymphaeaceae) alkaloids in leaves or seeds are well known. However, the sleep-promoting activity of the lotus rhizome (LE), which is used mainly as food, has not yet been evaluated.ObjectiveWe investigated the sleep-promoting activity of LE water extract.Materials and methodsInstitute of Cancer Research (ICR) mice (n = 8) were subject to a pentobarbital-induced sleep test to assess changes in sleep latency and duration following the administration of LE (80–150 mg/kg). In addition, electroencephalography analysis was performed to determine the sleep quality after LE treatment as well as the sleep recovery effect of LE using a caffeine-induced insomnia SD rat model. Real-time PCR and western blot analysis were performed to investigate the expression of neurotransmitter receptors, and the GABA_A receptor antagonists were used for receptor binding analysis.ResultsAn oral administration of 150 mg/kg LE significantly increased sleep duration by 24% compared to the control. Furthermore, LE increased nonrapid eye movement (NREM) sleep by increasing theta and delta powers. In the insomnia model, LE increased sleep time by increasing NREM sleep. Moreover, treatment with picrotoxin and flumazenil decreased the sleep time by 33% and 23%, respectively, indicating an involvement of the GABA_A receptor in the sleep-enhancing activity of LE. The expression of GABA_A receptors and the concentration of GABA in the brain were increased by LE.Discussion and conclusionsThe results suggest that the sleep-promoting activity of LE was via the GABA_A receptor. Collectively, these data show that LE may promote sleep.     

Ⅲ型胶原α链基因mRNA的表达与舌苔形成的关系研究

目的研究Ⅲ型胶原α链基因表达水平与舌苔形成之间的关系。方法应用基因芯片技术和实时荧光定量逆转录-聚合酶链反应（RT-PCR）技术,检测Ⅲ型胶原α链在不同舌苔中的表达水平。结果基因芯片显示,常见舌苔中Ⅲ型胶原α链表达水平差异有统计学意义;实时荧光定量PCR证实从高到低依次为：黄厚苔〉白薄苔〉白厚苔〉无苔〉黄薄苔〉胎儿白薄苔。结论常见舌苔中,Ⅲ型胶原α链表达水平有差异,提示Ⅲ型胶原α链基因表达水平的调控在舌苔形成过程中可能有一定的意义。