[Three-dimensional analysis of force distribution on maxillary dentition while distalization of first and second molars simultaneously with clear aligners].

Objective: To explore the force distribution on the maxillary dentition when the first and second molars distalized simultaneously with different step sizes using clear aligners in vitro in order to provide a theoretical basis for the rational design of molar distalization. Methods: Clear aligners were designed to simultaneously distalize the maxillary first and second molars bilaterally, with rectangular attachments placed on the buccal surfaces of the first and second premolars, as well as the second molars. Based on different step sizes, the aligners were divided into three groups: Group A (0.15 mm per step), Group B (0.20 mm per step), and Group C (0.25 mm per step). Ten aligners were fabricated for each group using 0.76 mm thick polyethylene terephthalate glycol (PET-G) sheets. A three-dimensional force measurement system was used to measure the forces exerted on each tooth by the aligners, the first and second molars served as the target teeth and the remaining teeth as anchorage teeth. The three-dimensional force data were compared among the three groups. Results: In the mesiodistal direction, the forces on the central and lateral incisors were relatively small among all three groups, with no statistically significant differences (P>0.05). However, significant differences were observed in the forces on the canines, first premolars, second premolars, first molars, and second molars (P<0.05). The distal forces on the second molars in Groups B and C were (6.13±1.45) N and (6.83±1.58) N, respectively, significantly higher than that in Group A [(3.51±1.01) N] (P<0.05). The distal force on the first molars in Group C [(6.62±0.89) N] was significantly higher than that in Groups A and B (P<0.05). The mesial reactive forces on the first and second premolars in Groups B and C were significantly higher than those in Group A (P<0.05). The mesial reactive force on the canines in Group C [(-2.98±1.33) N] was significantly higher than that in Group A [(-1.69±0.68) N] (P<0.05), while there were no significant differences between Groups B and C in the forces on the canines, first premolars, and second premolars (P>0.05). In the buccolingual direction, there were no statistically significant differences in the forces on the central and lateral incisors among three groups (P>0.05), but significant differences were observed in the forces on the canines, second premolars, and second molars (P<0.05). The buccolingual forces on the canines, second premolars, and second molars in Group B were (-0.56±0.54), (-2.07±0.95), (1.13±0.55) N, respectively, significantly higher than those in Group A (P<0.05), but there were no significant differences compared to Group C (P>0.05). Compared to the mesiodistal and buccolingual forces, the vertical forces on the target and anchorage teeth were relatively small in all three groups. Conclusions: When using 0.76 mm thick PET-G sheets to fabricate clear aligners for simultaneous molar distalization, a step size of 0.20 mm per step is recommended. To prevent buccal tipping of the molars during distalization, it is advisable to design lingual displacement for the molars and buccal displacement for the adjacent anchorage teeth to counteract the unfavorable forces, with attachments placed on the primary anchorage teeth.

[Research advances of collagen-based biomaterials in wound repair].

Acute or chronic wounds are common clinical problems. Collagen, with advantages including rich sources, impeccable biocompatibility, and inherent biodegradability, has been widely used in fundamental research and clinical treatment of wound repair with broad prospects of clinical applications. This article provided a brief overview of the role of collagen in various biological processes related to wound healing and also outlined the sources of collagen. Furthermore, the article summarized the application and recent research advancements of collagen-based wound dressings in the field of wound repair.

[shRNA-Mediated Suppression of γ-Synuclein Leading to Downregulation of p38/ERK/JNK Phosphorylation and Cell Cycle Arrest in Endometrial Cancer Cells].

In this study, we explored the effects of treating human endometrial cancer cells with γ-synuclein-specific short hairpin RNA (shRNA) and elucidated the associated mechanisms in vitro and in vivo through the p38, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK) signaling pathways. Cell proliferation and migration were assessed using CCK8, Transwell, and scratch wound healing assays. Flow cytometry and laser scanning confocal microscopy were used to detect cell cycle changes. Relative levels of phosphorylated and non-phosphorylated (p) p38, ERK1/2 and JNK1/2/3 were determined in vitro and in vivo using simple western blotting assays. Cell proliferation in the experimental group decreased significantly and cells transfected with shRNA showed reduced migration rates (P < 0.05). p-p38, p-ERK1/2, and p-JNK1/2/3 levels were downregulated in the experimental group in vitro and in vivo. Tumor volumes and weights in the experimental group were significantly lower (P < 0.05). Tumor formation time in the negative control group was significantly shorter (P < 0.05). Flow cytometry showed that the number of cells in the G1 and mitotic phases increased and that in the S phase decreased after SNCG silencing (P < 0.05). Confocal microscopy showed that the percentage of cells in the mitotic phase increased after SNCG gene silencing (P < 0.05). We conclude that shRNA-mediated suppression of γ-synuclein decreased the proliferation, migration, and tumorigenicity of endometrial cancer cells via downregulation of p38, ERK, and JNK phosphorylation. High SNCG expression is closely related to the growth cycle of endometrial cancer cells.

[Current situation of initial female fertility evaluation in assisted reproductive institutions in China in 2019].

The evaluation of female fertility at first diagnosis is an important premise and basis for determining the treatment scheme of assisted reproduction. In this study, a survey was conducted among 13 assisted reproductive institutions in China to understand the evaluation indicators and detection methods of female fertility at first diagnosis in various institutions, and provide a basis for reasonable selection of indicators. The survey showed that the indicators of female fertility evaluation at first diagnosis among assisted reproductive institutions included general health indicators, ovarian reserve indicators, and uterine conditions, etc. The selection of indicators was considerably consistent, but the detection methods were quite different. Therefore, it is necessary to choose the detection method with better validity and less harm.

Assisted reproductive technology service availability, efficacy and safety in mainland China: 2016.

STUDY QUESTION: What is the current status of assisted reproductive technology (ART) service availability, efficacy and safety in mainland China? SUMMARY ANSWER: In this first national report on ART status in mainland China, data on treatment numbers, outcomes and complications in 2016 are provided and analyzed, respectively. WHAT IS KNOWN ALREADY: National ART Service Provision Surveys are conducted in mainland China regularly. Data were analyzed, and this manuscript was written by team members from the National Center for Women and Children's Health, China CDC and the Department of Women and Children Health, National Health Commission of the People's Republic of China. STUDY DESIGN, SIZE AND DURATION: A cross-sectional nationwide survey was completed in 2018, in which data regarding ART treatments, performed from 1st January to 31st December2016 in 445 ART clinics located in 31 provinces of mainland China, were collected. PARTICIPANTS/MATERIALS, SETTING AND METHODS: There were in total 451 licensed ART clinics (including artificial insemination clinics) in mainland China in 2016, of which 445 submitted service data. A total of 906 840 cycles were provided by 323 in vitro fertilization (IVF) clinics, involving 375 770 conventional IVF cycles, 154 948 intracytoplasmic sperm injection (ICSI) cycles, 367 146 frozen embryo transfer (FET) thawing cycles and 8976 preimplantation genetic diagnosis (PGD) treatment cycles. A total of 161 376 artificial (i.e. intrauterine) insemination (AI) cycles were reported by 443 clinics, with 126 872 cycles using the husband's semen (AIH) and 34 504 using donor semen (AID). MAIN RESULTS AND THE ROLE OF CHANCE: In total, 98.7% of the licensed clinics, contributing to 100% of the ART services (including AID and AIH cycles), were included in this report. (Six clinics provided institutional information only and were excluded.) There were 906 840 in vitro fertilization cycles performed in mainland China with a population of over 1.3 billion inhabitants, with cycles per million inhabitants (C/M) increasing from 360 in 2013 to 657 in 2016, nationwide (range among provinces: 45-3676). After treatment with conventional IVF, the clinical pregnancy rate (PR) per oocyte retrieval cycle was 23.2%, the delivery rate (DR) per oocyte retrieval cycle was 18.7% and the proportion of twin delivery among the total deliveries was 27.9%. For ICSI cycles, the PR, DR and TDR were 20.5%, 16.7% and 27.2%, respectively. For FET per thawing cycles, the PR, DR and TDR were 48.2%, 37.6% and 24.2%. For PGD per diagnosis cycles, the PR, DR and TDR were 38.1%, 29.7% and 4.2%. For AIH cycles, the PR and DR were 13.3% and 10.5%; for AID cycles, the PR and DR were 24.3% and 21.1%, respectively. The total number of live infants born in mainland China in 2016, was 18.46 million, and the number of infants born through ART conducted in 2016 was 311 309, which accounted for 1.69% of the total. The reported rate of birth defects was about 87/10 000. The incidence of moderate to severe ovarian hyper-stimulation syndrome (OHSS) was 11.5 per 1000 oocyte retrieval cycles, and other complications were much more rare. LIMITATIONS AND REASONS FOR CAUTION: This report is based on the summary data of ART services provided. The success rates were not calculated by age stratification. A low rate of birth defects was reported, which might be confounded by variations in birth follow-up methods, statistical timing and record taking. WIDER IMPLICATIONS OF THE FINDINGS: ART service availability has improved significantly in recent years in mainland China. Because China is a vast country, significant imbalances in ART service provision do exist; however, the main efficacy and safety indicators were close to those of western countries. TRIAL REGISTRATION NUMBER: N/A. STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the National Key R&D Program of China (2016YFC1000307-2). There are no competing interests.

[Clinical, molecular genetic analysis, and treatment of 3 children with sitosterolemia].

Objective: To investigate clinical, molecular genetic characteristics, and treatment outcomes of 3 children with sitosterolemia. Methods: Three cases of children presented with multiple xanthomas during June 2016 to June 2017 were included. The clinical manifestations, laboratory examinations and follow-up data were retrospectively analyzed. DNA was extracted from peripheral blood and analyzed with whole exome sequencing(WES). All the detected variants were confirmed by Sanger sequencing. Plasma plant sterol concentrations were measured by gas chromatography-mass spectrometry. Results: Three cases of children including 1 boy and 2 girls presented with multiple linear and intertriginous xanthomas around skin of the joint areas at the age from 15 months to 6 years and 2 months. Total cholesterol of the 3 cases was elevated to 14.45, 15.47 and 15.85 mmol/L (3.36-6.46), and low density lipoprotein cholesterol was 9.02, 13.54 and 12.47 mmol/L (< 3.36) respectively. Genetic analysis with WES revealed that 2 cases carried compound heterozygous variants in ABCG5 gene, 1 case carried compound heterozygous variants in ABCG8 gene. Two reported variants (p. N437K, p.R446X) and one novel variant (p.Q251X) of ABCG5 were identified in case 2 and 3. Two novel ABCG8 variants (p.R263Q, c.1528_1530delATC) were found in case 1. All these children had extremely high plasma plant sterol levels, thus the diagnosis of sitosterolemia was confirming. The campesterol level was 111.35, 102.86 and 58.91 µmol/L(0.01-10.00), the stigmasterol was 14.97, 29.43 and 17.79 µmol/L (0.10-8.50) and the sitosterol was 231.20, 177.66 and 114.20 µmol/L (1.00-15.00) respectively. The total serum cholesterol levels of three children decreased to nomal after the patients were placed on the low plant sterol/low cholesterol diet. The xanthomas regressed gradually, and almost disappeared after 8 months of treatment in case 1 and 3. Conclusions: Children with sitosterolemia presented with skin xanthomas around the joint areas. The level of total cholesterol, low density lipoprotein cholesterol and plant sterols increased obviously. One novel variant (p.Q251X) of ABCG5 and 2 novel variants (p.R263Q, c.1528_1530delATC) of ABCG8 were identified. Children with sitosterolemia responded well to a low plant sterols/low cholesterol diet.

Association of ectomycorrhizal fungi with Picea crassifolia (Pinaceae, Piceoidae) from high-altitude stands in Mount Helan Nature Reserve, China.

We investigated the diversity of ectomycorrhiza associated with the endemic Picea crassifolia in Mount Helan National Nature Reserve in Inner Mongolia, China. Toward this objective, we conducted morphological and molecular identification of ectomycorrhizae in soil cubes taken from pure P. crassifolia stands. Eleven types of ectomycorrhizal (ECM) organisms were separated, briefly described, and identified. Nine morphotypes belonged to the phylum Basidiomycotina [Amphinema byssoides, Cortinarius sp (cf. limonius), Cortinarius vernus, Inocybe cf. nitidiscula, Inocybe sp 1, Sebacina incrustans, Sebacina sp, Suillus luteus, and Piceirhiza tuberculata x Picea crassifolia (comb. Nov.)], and two morphotypes to the phylum Ascomycotina (Cenococcum geophilum and Helvella sp). The diversity of ECM organisms in P. crassifolia was lower than that reported by other studies on spruce or pine forests, or on sporocarp diversity in the high-mountain forests of China. Most of the fungi in the rhizosphere did not correspond to species previously recorded as sporocarps above ground. Here, several new ectomycorrhiza morphotypes are proposed and described. We also confirmed the ectomycorrhizal status of the genus Sebacina (order Sebacinales).

The clinical significance of CA19-9 in ovarian mature cystic teratoma.

OBJECTIVE: To evaluate the clinical significance of CA19-9 in patients with ovarian mature cystic teratoma (MCT). MATERIALS AND METHODS: A retrospective study was performed on 65 patients with pathologically-confirmed MCT and 80 patients with benign epithelial ovarian tumors. Serum tumor markers for all patients and tissue CA19-9 for MCTs were measured. The relationships between clinical characteris- tics of MCTs and CA19-9, as well as the correlation between serum and tissue level of CA19-9 in MCTs, were evaluated. RESULTS: The mean serum level of CA19-9 in MCTs was significantly higher than that in benign ovarian epithelial tumors (49.9 ± 73.4 IU/ml vs. 17.08 ± 24.8 IU/ml). CA19-9 was the only tumor marker with a mean serum level above the cut-off value and the elevation rate was 30.76% in MCTs. The positive tissue expression rate of CA19-9 in MCT patients were 50.9% and were higher than that of preoperative serum levels (50.9% vs. 32.7%). CONCLUSION: Serum CA19-9 has the highest positivity rate among other tumor markers in MCT. Elevated serum CA19-9 is not an uncommon finding MCT and could be used as a marker in the differential diagnosis of MCT in patients with pelvic mass.

Enhanced effect of ß-tricalcium phosphate phase on neovascularization of porous calcium phosphate ceramics: in vitro and in vivo evidence.

Neovascularization plays a key role in bone repair and regeneration. In the present study, four types of porous calcium phosphate (CaP) ceramics, namely hydroxyapatite (HA), biphasic calcium phosphates (BCP-1 and BCP-2) and ß-tricalcium phosphate (ß-TCP), with HA to ß-TCP ratios of 100/0, 70/30, 30/70 and 2/98, respectively, were investigated in terms of their angiogenic induction. The in vitro cell culture revealed that the ceramics could promote proliferation and angiogenesis of human umbilical vein endothelial cells (HUVECs). This result could be achieved by stimulating CCD-18Co human fibroblasts to secrete angiogenic factors (vascular endothelial growth factor, basic fibroblast growth factor and transforming growth factor-ß) as a paracrine effect, as well as by up-regulating HUVECs to express these angiogenic factors and their receptors (KDR, FGFR1 and ACVRL1) and the downstream eNOS as an autocrine effect. These effects were more significant in ß-TCP and BCP-2, which had a higher content of ß-TCP phase. In the in vivo implantation into the thigh muscles of mice, the process of neovascularization of the ceramics was initiated at 2 weeks and the mature vascular networks were formed at 4 weeks as visualized by hematoxylin and eosin staining and scanning electron microscopy. Microvessel density count confirmed that ß-TCP and BCP-2 induced more microvessels to form than HA or BCP-1. This phenomenon was further confirmed by the significantly up-regulated expressions of angiogenesis-related genes in the ingrowth of cells into the inner pores of the two ceramics. All the results confirmed the angiogenic induction of porous CaP ceramics, and a higher content of ß-TCP phase had an enhanced effect on the neovascularization of the ceramics.

Enhanced repair of a critical-sized segmental bone defect in rabbit femur by surface microstructured porous titanium.

Repair of load-bearing bone defects remains a challenge in the field of orthopaedic surgery. In the current study, a surface microstructured porous titanium (STPT) successively treated with H2O2/TaCl5 solution and simulated body fluid was used to repair the critical-sized segmental bone defects in rabbit femur, and non-treated porous titanium (NTPT) and porous biphasic calcium phosphate ceramics (PBCP) were used as control, respectively. A 15 mm long implant was positioned in the femoral defect and stabilized by a plate and screws fixation. After implantation into the body for 1, 3 and 6 months, X-ray observation confirmed that porous titanium groups (NTPT and STPT) provided better mechanical support than PBCP group at the early stage. However, there was no obvious difference in the formed bony callus between PBCP and STPT groups in the later stage, and they both showed better shape of bony callus than NTPT group. Micro-CT and histomorphometric analysis for the samples of 6-month implantation demonstrated that more new bone formed in the inner pores of PBCP and STPT groups than that in NTPT group. Moreover, the biomechanical tests revealed that STPT group could bear larger compressive load than NTPT and PBCP groups, almost reaching the level of the normal rabbit femur. STPT exhibited the enhanced repairing effect on the critical-sized segmental bone defect in rabbit femur, meaning that it could be an ideal material for the repair of large bone defect in load-bearing site.

The enhanced effect of surface microstructured porous titanium on adhesion and osteoblastic differentiation of mesenchymal stem cells.

Porous titanium with appropriate surface treatments can be osteoinductive. To investigate the effect of surface treatments of porous titanium on the attachment and differentiation of mesenchymal stem cells (MSCs), two kinds of surface microstructured porous titaniums, H2O2/TaCl5 treated one (HTPT), and H2O2/TaCl5 and subsequent simulated body fluid (SBF) treated one (STPT) were fabricated, and non-treated one (NTPT) was used as control. The morphology, specific surface area (SSA), pore distribution and mechanical strength of these materials were characterized respectively, and the results showed that H2O2/TaCl5 treatment led to a significant increase in both SSA and micropores of HTPT, and the further SBF immersion resulted in the formation of a layer of bone-like apatite on the surface of STPT. Although the surface treatments had a little negative impact on the compressive strength and elasticity modulus of porous titanium, the mechanical strength of HTPT or STPT was enough for the bone defect repair of the load-bearing sites. The protein adsorption and cell adhesion experiments confirmed that the microstructured surface notably enhanced porous titanium's protein binding capacity and promoted MSCs adhesion on the surface. More importantly, cell differentiation experiments proved that the microstructured surface evidently elevated the osteoblastic gene expressions of MSCs compared to NTPT. The enhanced biological effect by the surface treatments was more robust on STPT. Therefore, our results suggest that the microstructured surface has great potential for promoting MSCs differentiation towards osteoblasts, giving excellent support for the osteoinduction of porous titanium with appropriate surface treatments.

First Report of Rice stripe virus in Vietnam.

Rice stripe virus (RSV) is the type member of the genus Tenuivirus, and one of the prevalent viruses infecting rice. The disease was first recorded in central Japan in 1903, and is currently present in many Asian countries, including South Korea and China in the Far East (1,2). In May of 2012, a disease outbreak in Indica rice (Oryza satira L.) caused losses in a field in Huaifu, Hungyen, Vietnam (20°53'N, 106°02'E). Infected plants showed yellowing stripe symptoms on leaves. A survey indicated that disease incidence was about 10%. Six leaf samples were randomly collected and four were found positive in dot-ELISA using RSV-specific monoclonal antibodies (provided by Dr. X. Zhou, Institute of Biotechnology, Zhejiang University) (3). To confirm RSV infection, total RNA was extracted from dot-ELISA positive and asymptomatic control samples. RT-PCR was performed using RSV-specific primers (CP(+): 5'-GAGGATCCATGGGTACCAACAAGCCAG-3', CP(-): 5'-TCGTCGACCTAGTCATCTGCACCTTCTG-3'; SP(+): 5'-TGGGATCCATGCAAGACGTACAAAGGAC-3', SP(-): 5'-CTGTCGACCTATGTTTTATGAAGAAGAGGT-3'; NS2(+): 5'-GAGGATCCATGGGTACCAACAAGCCAG-3', NS2(-): 5'-CCGTCGACTCATACATCTGAATTTG-3'; NS3(+): 5'-ACCGGATCCATGACTATCAAATACAAC-3', NS3(-): 5'-CCGTCGACTCATACATTAGCTATTGTC-3') that amplify the coat protein, disease-specific protein, and NS2 and NS3 genes of RSV, respectively. Amplicons of the expected size were obtained from the four symptomatic but not the asymptomatic plants. Amplicons obtained from one of the positive samples were cloned into the vector pMD18-T (TaKaRa, Dalian, China) and sequenced (GenBank Accession Nos. KC197055 to KC197058). Sequence comparisons indicated that the complete sequences of the CP, SP, NS2, and NS3 of the Vietnamese isolate shared 97.1%, 97.5%, 96.8%, and 97.3% sequence identity at the nucleotide level with the corresponding genes of RSV isolate T (NC_003776, NC_003753, and NC_003754, respectively). These results indicate that the virus associated with yellowing stripe disease of rice in Vietnam is an isolate of RSV. To our knowledge, this is the first report of RSV in Vietnam. This finding redefined the distribution of RSV in the world. Research on whole-genome sequencing of the Vietnamese isolate is continuing and is being expanded to compare the genetic diversity of the virus, assisting in the study of the evolution of the virus. References: (1) S. Toriyama. Microbiol. Sci. 3:347, 1986. (2) Q. Y. Lin et al. J. Fujian. Agric. Univ. 20:24, 1991. (3) G. Z. Wang et al. Acta Phytopathol. Sinica 34:302, 2004.

Three dimensional microfluidics with embedded microball lenses for parallel and high throughput multicolor fluorescence detection.

We report a 3D microfluidic device with 32 detection channels and 64 sheath flow channels and embedded microball lens array for high throughput multicolor fluorescence detection. A throughput of 358 400 cells/s has been accomplished. This device is realized by utilizing solid immersion micro ball lens arrays for high sensitivity and parallel fluorescence detection. High refractive index micro ball lenses (n = 2.1) are embedded underneath PDMS channels close to cell detection zones in channels. This design permits patterning high N.A. micro ball lenses in a compact fashion for parallel fluorescence detection on a small footprint device. This device also utilizes 3D microfluidic fabrication to address fluid routing issues in two-dimensional parallel sheath focusing and allows simultaneous pumping of 32 sample channels and 64 sheath flow channels with only two inlets.

Chondrogenic differentiation of mesenchymal stem cells induced by collagen-based hydrogel: an in vivo study.

Chondrogenic differentiation of mesenchymal stem cells (MSCs) relies on inductive media of chondrogenic environment. With proper design, a cellular microenvironment mimicking chondrogenic environment might be created to induce chondrogenesis of MSCs. In this study, bone marrow mesenchymal cells (BMSCs) were encapsulated in collagen-based hydrogel, and then enclosed in diffusion-chambers which allow the body fluid to permeate and preclude the host cells to invade. Then, the chamber with the hydrogel-BMSCs composite was implanted in the back of rabbits subcutaneously. The specimens in the chamber were harvested for histological, immunohistochemical, and RT-PCR analyses after 8 weeks. The results showed that cells with the characteristic of chondrocytes were homogenously distributed and the extracellular matrix (ECM) of cartilage has been secreted, indicating the chondrogenic differentiation of BMSCs. As control, nothing was obtained with only BMSCs. Moreover, the expression of collagen type II, indicator of cartilage ECM, was less in tissues with collagen-alginate-hydrogel (CAH) than that with collagen-hydrogel (CH). The results showed that both CH and CAH may induce the chondrogenesis and the induction is materials dependent. From in vitro experiments, TGF-beta is a necessary signal molecule for chondrogenesis, and it was suggested that the material may take in vivo growth factors to trigger chondrogenesis. From the studies, the chondrogenic induction of the hydrogel may be ascribed to that the hydrogel may provide a suitable environment and aggregate the signal molecule for chondrogenesis in vivo. The results would lend valuable reference in clinical for selection of appropriate scaffold for cartilage repair.

Genetic polymorphisms in glutathione S-transferases T1, M1 and P1 and susceptibility to reflux esophagitis.

Recent studies indicate that the prevalence of reflux esophagitis (RE) in China is increasing. RE is one of the most common esophageal complications associated with gastroesophageal reflux disease (GERD) and RE-Barrett's esophagus-esophageal adenocarcinoma (EAC) sequence has been considered as an histogenesis model for EAC in Western countries. RE is only present in a subset of patients with GERD, suggesting an altered susceptibility to RE may exist in these GERD individuals. However, the genetic changes related with high susceptibility to RE is largely unknown. The polymorphisms in glutathione S-transferases (GSTs) T1, M1 and P1 have been reported with high susceptibity to esophageal cancer in Chinese people. The present case-control study was thus undertaken to characterize the genetic polymorphisms of GSTs and their correlation with susceptibility to RE. One hundred and nine patients with RE, 97 patients with nonerosive reflux disease (NERD) and 97 normal controls were recruited in this study. All the subjects were from Beijing, China, and received endoscopic examination and questionnaires for RE. Genomic DNA was extracted from the lymphocytes of peripheral blood for each subject. Genotypes of the GSTM1 and GSTT1 genes were analyzed by a multiplex PCR method. A-->G polymorphism of codon 104 of the GSTP1 gene was detected using PCR-based restriction fragment length polymorphisms (RFLP). The variant GSTP1 genotypes (*A/*Bomicron*B/*B) was found with a high frequency in the case with RE (40%), and followed by NERD (25%) and normal control (22%). The differences were statistically significant (P < 0.05). The risk for RE increased 2.42-fold [odds ratio (OR); 95% confidence interval (95% CI), 2.42 (1.22-4.80)] in the subjects with variant GSTP1 genotype. The subjects with positive variant GSTP1 genotypes and negative H. pylori infection showed increasing tendency for risk of RE [OR (95% CI), 2.67 (1.06-6.70)]. However, the subjects with GSTT1 and GSTM1 polymorphisms did not show any correlation with high risk for RE or NERD. No significant interactions were identified between the variant GSTs and cigarette smoking, or alcohol drinking and subtype of RE. The present result suggests that GSTP1 genetic polymorphism may be one of the high susceptibility factors involved in the mechanisms of RE. H. pylori infection may play a protective role against RE.

Induction of a germline mutation at a hypervariable mouse minisatellite locus by 252Cf radiation.

Male C3H/HeN mice were exposed to 252Cf radiation and mated with unirradiated C57BL/6N females. F1 mice were analyzed for germline mutation at the paternally derived C3H/HeN allele of a hypervariable minisatellite locus, Ms6hm. This locus exhibited a high frequency of length change mutation spontaneously, and the mutation frequency of the paternally derived C3H/He allele in F1 mice born to unirradiated males was 8.4%. Exposure of male mice to 252Cf radiation resulted in even higher frequency of germline mutation. The spermatid stage germ cells were most sensitive to neutrons, and the mutation frequencies of the paternal allele were elevated to 18%, 26% and 24% for 0.35, 0.7 and 1.02 Gy of 252Cf radiation, respectively. Spermatozoa and spermatogonia stages were less sensitive and the mutation frequencies for 1.02 Gy of 252Cf radiation were 16% and 19%, respectively. The 252Cf radiation consisted of 35% gamma-rays and 65% neutrons. Assuming that these two radiations act additively, RBE of 252Cf neutrons for the induction of minisatellite mutation was calculated to be 5.9 for spermatozoa stage irradiation, 2.6 for spermatid stage irradiation and 6.5 for spermatogonia irradiation.

Association of minisatellite instability with c-myc amplification and K-ras mutation in methylcholanthrene-induced mouse sarcomas.

Instability of microsatellite sequences are frequently found in human tumors. In addition, minisatellite sequences, another group of highly unstable sequences, serve as sensitive markers of genetic instability. We have studied minisatellite instability in methylcholanthrene-induced mouse sarcomas. These sarcomas frequently carry the amplified c-myc gene. Seven sarcomas without the amplification and seven others with the amplification were selected randomly. Regardless of the state of the c-myc gene amplification, these sarcomas exhibited a varying degree of transplantability in syngeneic mice. The hypervariable mouse minisatellite locus Ms6hm was found to be highly unstable, specifically among sarcomas with the amplified c-myc gene. However, chromosome instability, as analyzed by micronucleus assay, was observed similarly for two groups of sarcomas. In addition, transversion of G to C and A to T was detected at the K-ras gene in four of the seven sarcomas with the amplified c-myc gene, and these mutations are thought to be induced directly by methylcholanthrene. Thus, concomitant occurrence was observed for three seemingly unrelated mutations, amplification of the c-myc locus, point mutation of the K-ras gene, and instability at the hypervariable mouse minisatellite locus. The present study indicates a possible involvement of K-ras mutation and c-myc amplification in induction of genetic instability in methylcholanthrene-induced mouse sarcomas.