Online prediction model for primary aldosteronism in patients with hypertension in Chinese population: A two-center retrospective study.

Background: The prevalence of primary aldosteronism (PA) varies from 5% to 20% in patients with hypertension but is largely underdiagnosed. Expanding screening for PA to all patients with hypertension to improve diagnostic efficiency is needed. A novel and portable prediction tool that can expand screening for PA is highly desirable. Methods: Clinical characteristics and laboratory data of 1,314 patients with hypertension were collected for modeling and randomly divided into a training cohort (919 of 1,314, 70%) and an internal validation cohort (395 of 1,314, 30%). Additionally, an external dataset (n = 285) was used for model validation. Machine learning algorithms were applied to develop a discriminant model. Sensitivity, specificity, and accuracy were used to evaluate the performance of the model. Results: Seven independent risk factors for predicting PA were identified, including age, sex, hypokalemia, serum sodium, serum sodium-to-potassium ratio, anion gap, and alkaline urine. The prediction model showed sufficient predictive accuracy, with area under the curve (AUC) values of 0.839 (95% CI: 0.81-0.87), 0.814 (95% CI: 0.77-0.86), and 0.839 (95% CI: 0.79-0.89) in the training set, internal validation, and external validation set, respectively. The calibration curves exhibited good agreement between the predictive risk of the model and the actual risk. An online prediction model was developed to make the model more portable to use. Conclusion: The online prediction model we constructed using conventional clinical characteristics and laboratory tests is portable and reliable. This allowed it to be widely used not only in the hospital but also in community health service centers and may help to improve the diagnostic efficiency of PA.

The CYP3A5 and ABCB1 Gene Polymorphisms in Kidney Transplant Patients and Establishment of Initial Daily Tacrolimus Dosing Formula.

BACKGROUND: Tacrolimus is an immunosuppressive drug used to prevent organ rejections. Many factors could influence blood concentration of tacrolimus. OBJECTIVE: To detect genotypes of cytochrome P450 3A5 (CYP3A5) and ABCB1 in kidney transplant patients and establish initial daily tacrolimus dosing formula based on genotypes of CYP3A5 and ABCB1 and patients' clinical parameters. METHODS: Sequence specific primer polymerase chain reaction (PCR) and PCR restriction fragment length polymorphism were used to detect genotypes of CYP3A5 and ABCB1. The blood cell, procalcitonin, C-reactive protein, height, weight, age, gender and other clinical parameters were recorded. Multiple linear regression analysis and Pearson correlation analysis were used to conduct date analysis. RESULTS: 102 cases were enrolled in cohort 1, and there were 10 cases of CYP3A5 *1/*1 (9.8%), 28 cases of CYP3A5 *1/*3 (27.5%), and 64 cases of CYP3A5 *3/*3 (62.7%). The distributions of ABCB1 C3435T genotype were CC 36 (35.3%), CT 52 (51.0%), and TT 14 (13.7%). The distributions of ABCB1 G2677T/A genotype were GG 39 (38.2%), GT 40 (39.2%), and TT 23 (22.5%). The formula was 7.499 + (0.053 × Weight) - (0.029 × Hemoglobin concentration) - (1.045 × CYP3A5 genotype) (CYP3A5 genotype: *1/*1 type inputs 0, *1/*3 type inputs 1, *3/*3 type inputs 2). The predicted doses from the established formula had a significant correlation (r = 0.605) with actual clinical doses (P < 0.05). CONCLUSION AND RELEVANCE: Hemoglobin concentration, weight, and CYP3A5 genotype should be considered using tacrolimus. The initial daily tacrolimus dosing formula established can make a good prediction.

Activation of Gonadotropin-releasing Hormone Receptor Impedes the Immunosuppressive Activity of Decidual Regulatory T Cells via Deactivating the Mechanistic Target of Rapamycin Signaling.

Understanding maternal immune tolerance is crucial for the development of therapeutics for immunological pregnancy complications. Decidual regulatory T cells (Tregs) play a pivotal role in the maintenance of maternal immune tolerance. Using a murine allogeneic pregnancy model in the current study, we identified the up-regulation of gonadotropin-releasing hormone receptor (GnRHR) in decidual T cell subsets including CD4+ conventional T cells, CD8+ T cells, and CD4+Foxp3+ Tregs. Using a lentivirus-mediated GnRHR overexpression system and a GnRHR agonist, we found that GnRHR activation decreased the expression of Treg functional molecules such as IL10 (IL-10), IL-35 subunit EBI3 (Ebi3), IL2RA (CD25), TNFRSF18 (GITR), ICOS, and Treg master regulator FOXP3. The functional analysis indicated that GnRHR activation impairs the ability of Tregs to inhibit conventional T cell proliferation. We also revealed that GnRHR activation suppressed the mechanistic target of rapamycin (mTOR) signaling in GnRHR-overexpressing splenic Tregs (Wild type C57BL/6 J background) and decidual Tregs. MHY1485, a potent mTOR activator, effectively abolished the effect of the GnRHR agonist and promoted the immunosuppressive capability of Tregs. Furthermore, in an adoptive transfer model, Treg-specific GnRHR knockdown increased Foxp3 expression in decidual Tregs while decreasing the production of IFN-γ and IL-17 in decidual effector CD4+ T cells and reducing the production of IFN-γ in decidual effector CD8+ T cells. Taken together, the present study unveils a novel mechanism by which the immunosuppressive function of decidual Tregs is modulated, and deepens our understanding of maternal immune tolerance.

Prenatal Genetic Counseling in a Chinese Pregnant Woman With Rare Thalassemia: A Case Report.

Background: Prenatal genetic counseling can be difficult, especially when it is related to fetuses with a rare thalassemia. An intronic variant located far from obvious regulatory sequences in the HBB gene could be very difficult to evaluate as it may affect the mRNA processing or cause ß-thalassemia (ß-thal). In the present study, a Chinese pregnant woman with HbJ-Bangkok and a very rare change in the second intron of the HBB gene [IVS-II-806(G>C), NM_000518.4, HBB: c.316-45G>C] in combination with α+-thalassemia was reported, which can assist in prenatal genetic counseling. Case Report: A 26-year-old pregnant woman presented at the obstetric clinic for a routine pregnancy check at 12 weeks of gestation. Red blood counts and high-performance liquid chromatography (HPLC) were consistent with clinical manifestations of anemia. Multiplex gap-polymerase chain (gap-PCR) displayed rightward deletion (-α3.7/αα). Direct DNA sequencing of the δ-globin gene showed no mutation. Sanger sequencing of the ß-globin gene showed a previously undescribed condition of double heterozygosity for HbJ-Bangkok and a very rare change in the second intron of the HBB gene [IVS-II-806(G>C), NM_000518.4, HBB: c.316-45G>C] that has not been previously reported in the HbVar database. Thus, a rare combination of α+-thal and a compound heterozygosity of HbJ-Bangkok and [IVS-II-806(G>C)] with α+-thal (-α3.7/αα) was finally diagnosed. Prenatal genetic counseling was made based on the genotype and phenotype analyses. Conclusion: This study enlarges the mutation spectrum of ß-globin gene and emphasizes DNA analysis in resolving unusual patterns in Hb analysis and the importance of sharing the observed rare undefined mutations and the possible interactions with known molecular defects, which can assist in prenatal genetic counseling.

miR-5188 augments glioma growth, migration and invasion through an SP1-modulated FOXO1-PI3K/AKT-c-JUN-positive feedback circuit.

The biological effect and molecular mechanism of miR-5188 have not been thoroughly investigated. The study aims at elucidating the role of miR-5188 in glioma progression. Human glioma cell lines and tissues were used for functional and expression analysis. Cellular and molecular techniques were performed to explore the functions and mechanisms of miR-5188 in glioma. In our investigation, we demonstrated that miR-5188 promoted cell proliferation, the G1/S transition of the cell cycle, migration and invasion in glioma and reduced the lifespan of glioma-bearing mice. miR-5188 directly targeted FOXO1 and activated PI3K/AKT-c-JUN signalling, which enhanced miR-5188 expression. Moreover, the c-JUN transcription factor functionally bound to the miR-5188 promoter region, forming the positive feedback loop. The feedback loop promoted glioma progression through activating the PI3K/AKT signalling, and this loop is augmented by the interaction between SP1 and c-JUN. Moreover, it was also found that the miR-5188/FOXO1 axis is facilitated by SP1-activated PI3K/AKT/c-JUN signalling. In glioma samples, miR-5188 expression was found to be an unfavourable factor and was positively associated with the mRNA levels of SP1 and c-JUN, whereas negatively associated with the mRNA levels of FOXO1. Our investigation demonstrates that miR-5188 could function as a tumour promoter by directly targeting FOXO1 and participating in SP1-mediated promotion of cell growth and tumorigenesis in glioma.

Hypoglycemia during Short-Term Intensive Insulin Therapy and Its Association with Long-Term Glycemic Remission in Patients with Newly Diagnosed Type 2 Diabetes.

BACKGROUND: Short-term intensive insulin therapy induces long-term glycemic remission in half of patients with newly diagnosed type 2 diabetes. The concomitant hypoglycemia needs further analysis. METHODS: We collected data from three randomized trials conducted with the same inclusion and exclusion criteria at our institution from 2002 to 2015. Continuous subcutaneous insulin infusion (CSII) was provided to achieve the glycemic goals within a week and then maintained for 14 days. Hypoglycemia episodes during short-term treatment and the one-year drug-free glycemic remission were observed. RESULTS: A total of 244 patients were included. The per day episode of mild hypoglycemia (3.0-3.9 mmol/L) was higher in the remission group than in the nonremission group (0.26 ± 0.20 vs. 0.18 ± 0.21, P = 0.005). However, a moderate hypoglycemia episode (<3.0 mmol/L) per day was insignificantly lower in the remission group (0.02 ± 0.04 vs. 0.03 ± 0.04, P = 0.005). However, a moderate hypoglycemia episode (<3.0 mmol/L) per day was insignificantly lower in the remission group (0.02 ± 0.04 vs. 0.03 ± 0.04, P = 0.005). However, a moderate hypoglycemia episode (<3.0 mmol/L) per day was insignificantly lower in the remission group (0.02 ± 0.04 vs. 0.03 ± 0.04, P = 0.005). However, a moderate hypoglycemia episode (<3.0 mmol/L) per day was insignificantly lower in the remission group (0.02 ± 0.04 vs. 0.03 ± 0.04. CONCLUSIONS: Mild hypoglycemic episodes during the continuing insulin dose reduction period indicate a long-term drug-free euglycemic remission in patients with newly diagnosed type 2 diabetes. However, the insulin dosage should be reduced even more quickly in the future treatment to decrease the potential harms.

EXT1 and EXT2 Variants in 22 Chinese Families With Multiple Osteochondromas: Seven New Variants and Potentiation of Preimplantation Genetic Testing and Prenatal Diagnosis.

Multiple osteochondromas (MO), the most common type of benign bone tumor, is an autosomal dominant skeletal disorder characterized by multiple cartilage-capped bony protuberances. In most cases, EXT1 and EXT2, which encode glycosyltransferases involved in the biosynthesis of heparan sulfate, are the genes responsible. Here we describe the clinical, phenotypic and genetic characterization of MO in 22 unrelated Chinese families involving a total of 60 patients. Variant detection was performed by means of a battery of different techniques including Sanger sequencing and whole-exome sequencing (WES). The pathogenicity of the missense and splicing variants was explored by means of in silico prediction algorithms. Sixteen unique pathogenic variants, including 10 in the EXT1 gene and 6 in the EXT2 gene, were identified in 18 (82%) of the 22 families. Fourteen (88%) of the 16 variants were predicted to give rise to truncated proteins whereas the remaining two were missense. Seven variants were newly described here, further expanding the spectrum of MO-causing variants in the EXT1 and EXT2 genes. More importantly, the identification of causative variants allowed us to provide genetic counseling to 8 MO patients in terms either of preimplantation genetic testing (PGT) or prenatal diagnosis, thereby preventing the reoccurrence of MO in the corresponding families. This study is the first to report the successful implementation of PGT in MO families and describes the largest number of subjects undergoing prenatal diagnosis to date.

A new therapy in Epstein-Barr virus-associated lymphoproliferative disease: a case report and a revision of the literature.

BACKGROUND: Systemic chronic active Epstein-Barr virus infection is an extremely rare childhood disease. Since chronic active Epstein-Barr virus infection can trigger the onset of Epstein-Barr virus-associated lymphoproliferative disease. The clinical manifestations of the disease vary according to the site of involvement; therefore, management may be challenging. Currently, there are no standardized guidelines for treating Chronic active Epstein-Barr virus infection effectively. CASE PRESENTATION: We report a case of chronic active Epstein-Barr virus infection in a 5-year-old Chinese boy with intestinal, vascular, and neurological involvement. At age of 2 years and 7 months old, he had hepatomegaly and been diagnosed with Epstein-Barr virus infection. After treatment, he showed some clinical improvement. At age of 3 years and 3 months old, he presented with recurrent fever and diarrhea. Then he received methylprednisolone for 1 year and his symptoms ameliorated. At the age of 5 years, his symptoms recurred and had gastrointestinal hemorrhage and developed polyuria, frequent convulsions and hyponatremia. He was transferred to our hospital for further management. He was unconscious on admission and was diagnosised Epstein-Barr virus-lymphoproliferative disorder, based on the results in situ hybridization of EBV-encoded miRNA in sigmoid colon. Three-dimensional CT angiography demonstrated an aneurysm in the right internal carotid artery. Abdominal CT showed dilatation of vessels in part of the intestinal wall. He was also diagnosised Epstein-Barr virus encephalitis based on the elevated Epstein-Barr virus antibody titers and presence of Epstein-Barr virus DNA in the Cerebrospinal Fluid. A repeated duodenal artery embolization and symptomatic therapy could not control the hemorrhage after admission. He subsequently received treatment with ganciclovir, glucocorticoid, thalidomide, and propranolol. Hemorrhage was controlled in 5 days; his symptoms improved. The fever did not recur and the CSF pressure was also normalized. A follow-up CT at 3 months after admission showed regression of the aneurysm in the right internal carotid artery and the vascular lesion in the duodenum. DISCUSSION AND CONCLUSIONS: A new treatment protocol including thalidomide and propranolol resulted in a marked improvement in his clinical symptoms, and shows promise as a novel and effective therapeutic approach for Chronic active Epstein-Barr virus infection-associated lymphoproliferative disorder.

The phenomena of balanced effect between α-globin gene and of ß-globin gene.

BACKGROUND: Thalassemias (TM) are the most common autosomal recessive disorders in Southeast Asian countries. Both α- and ß-thalassemia lead to a decrease or absence of globin chains. The most serious of the thalassemia syndromes is thalassemia major which is characterized by a transfusion dependent anemia and subsequent iron overload caused by repeated blood transfusions. It is preventive by genotyping the parents. A better understanding of the laboratory data will help provide an accurate diagnosis of thalassemia major, and prevention and controlling programs in routine laboratories. CASE PRESENTATION: The patient was a one-year-old boy born to non-consanguineous parents. He was referred to our outpatient clinic for hemolytic anemia after a cold. Hematological investigations revealed severe anemia (Hb57 g/dL). The red cells displayed microcytosis, hypochromia and misshapen erythrocytes (MCV48.8 fL, MCH15.7 pg). Capillary electrophoresis (CE) electropherogram revealed normal level of HbA2 (3.2%) and elevated HbF (35.1%). The patient was diagnosed with ß-TM, based on severe microcytosis, hypochromia, normal Hb A2 and high Hb F level but no Hb H inclusion at the first visit. Later our molecular analysis revealed compound heterozygosity for codons 41-42 (-TTCT) (HBB: c.126_129delCTTT, ß0) and IVS-II-654 (C > T) (HBB: c.316-197C > T, ß+) mutation and deletional Hb H (--SEA/-α3.7). Thus, a combination of Hb H disease and a compound heterozygosity of ß+/ß0-thalassemia (ß+/ß0-thal) was finally diagnosed. CONCLUSIONS: Genotype-phenotype analysis shows that heterozygous mutations in the ß-globin gene could affect not only hematological parameters, but also elevate HbA2 levels. These effects could be ameliorated by the coinheritance of Hb H disease, which may be explained by the phenomena of the α-globin gene and of the ß-globin gene balanced effect.

A novel compound heterozygous GAA mutation in a Chinese family with juvenile onset form of Pompe disease with cardiomyopathy.

Pompe disease is an autosomal recessive disorder resulting from a deficiency of acid α-glucosidase (GAA). It is uncommon in the mainland of China, due to rare mutations in the GAA gene. The aim of this work was to elucidate the causative role of a novel compound heterozygous mutation of juvenile onset Pompe disease. In this study, clinical samples were obtained from two siblings with muscle weakness, recurrent airway infections, cardiomyopathy and respiratory insufficiency in a non-consanguineous Chinese family. The α-glucosidase activity in leukocytes of both children was low. Next-generation sequencing was performed on the 19 coding exons of GAA in both children, with confirmation by Sanger sequencing. Next-generation sequencing showed the same compound heterozygous GAA mutation (c.1216G>A p.Asp406Asn and c.1935C>A p.Asp645Glu) in both children. As this mutation is consistent with the clinical manifestations of juvenile onset Pompe disease and no other mutations were detected after scanning the gene sequence, we suggest that the Pompe disease phenotype is caused by compound heterozygosity for c.1216G>A and c.1935C>A. As c.1216G>A is not currently listed in the Pompe disease Mutation Database, this information about Pompe disease in a Chinese population is of particular interest.

[Analysis of hematological phenotype and genotype of 23 patients from Guangdong with co-inherited hemoglobin Hb Westmead and ß-thalassemia].

OBJECTIVE: To analyze the genotype-phenotype correlation among carriers from Guangdong with co-inherited hemoglobin Hb Westmead (HbWS) and ß-thalassemia. METHODS: Twenty three patients (including 9 males and 14 females, aged 1-53 year old) were diagnosed by hematological analysis and genetic testing. Complete blood cell count and hemoglobin electrophoresis analysis were performed on a XE4000i automatic hemocyte analyzer. Hb, HbF and HbA2 were tested by high performance liquid chromatography (HPLC). Gap-PCR was adopted to detect three common thalassemia deletions. Reverse dot-blotting (RDB) assay was applied for detecting three common non-deletional α2 gene mutations and ß-thalassemia. RESULTS: Among the 23 patients, 12 showed anemia, among whom 9 had mild anemia and 3 had moderate anemia. The lowest Hb was 68 g/L, and both mean corpuscular volume and mean corpuscular hemoglobin were lower than average, while HbA2 was higher than 3.5%. Genetic analysis confirmed that 5 cases had αWS-α/α-α, ß CD654/ß N (21.7%), 4 had α WS-α/α-α, ß CD41-42/ß N (17.4%), 5 had α WS-α/α-α, ß CD17/ß N (21.7%), 4 had α WS-α/α-α, ß CD28/ß N (17.4%), 1 had α WS-α/α-α, ß CD71-72/ß N (4.3%), 1 had αWS-α/α-α, ß CD27-28/ß N (4.3%), 1 had α WS-α/α-α, ß CD41-42/ß CD17 (4.3%), 2 had a concomitant ß-thalassemia heterozygosity and -α 3.7 deletion. CONCLUSION: Patients with co-existing Hb WS and other ß-thalassemia trait may show variable clinical features. Such compound heterozygotes are usually misdiagnosed during screening by hemoglobin electrophoresis, accurate diagnose should be attained by molecular diagnosis.

Sample Processing Effect on BK Virus Detection by Real-Time PCR in Urine Samples.

BACKGROUND: Measurement of BK viral DNA in urine by quantitative real-time PCR is a useful tool in monitoring BK viruria and nephropathy. We conducted this study to evaluate the differences in BK virus detection by different sample processing. METHODS: A total of 140 samples from 60 patients were processed by different methods including DNA extraction by spin column from whole urine, urine sediment, and urine supernatant. Boiled urine sediment lysate and untreated urine were also used in this study. RESULTS: The positive rate of BK virus was 34% for whole urine while other methods ranged from 21% - 27%. The quantification data showed that the maximum difference between whole urine and other methods varies from 2.36 - 2.47 log copies/mL in detection of BK virus load, while other methods showed minor differences. CONCLUSIONS: Our results highlight that centrifugation may cause BKV DNA to be lost in urine samples and whole urine is preferred for BK virus monitoring.

Dent Disease in Chinese Children and Findings from Heterozygous Mothers: Phenotypic Heterogeneity, Fetal Growth, and 10 Novel Mutations.

OBJECTIVE: To characterize the phenotypes of Dent disease in Chinese children and their heterozygous mothers and to establish genetic diagnoses. STUDY DESIGN: Using a modified protocol, we screened 1288 individuals with proteinuria. A diagnosis of Dent disease was established in 19 boys from 16 families by the presence of loss of function/deleterious mutations in CLCN5 or OCRL1. We also analyzed 16 available patients' mothers and examined their pregnancy records. RESULTS: We detected 14 loss of function/deleterious mutations of CLCN5 in 15 boys and 2 mutations of OCRL1 in 4 boys. Of the patients, 16 of 19 had been wrongly diagnosed with other diseases and 11 of 19 had incorrect or unnecessary treatment. None of the patients, but 6 of 14 mothers, had nephrocalcinosis or nephrolithiasis at diagnosis. Of the patients, 8 of 14 with Dent disease 1 were large for gestational age (>90th percentile); 8 of 15 (53.3%) had rickets. We also present predicted structural changes for 4 mutant proteins. CONCLUSIONS: Pediatric Dent disease often is misdiagnosed; genetic testing achieves a correct diagnosis. Nephrocalcinosis or nephrolithiasis may not be sensitive diagnostic criteria. We identified 10 novel mutations in CLCN5 and OCRL1. The possibility that altered CLCN5 function could affect fetal growth and a possible link between a high rate of rickets and low calcium intake are discussed.

Unexpected Detection of HbH-CS in a Pregnant Woman by Assessing HbA1c Using Capillary Electrophoresis.

BACKGROUND: Testing for hemoglobin A1C (HbA1C) is useful for following up on glycemic control in diabetic patients as well as in pregnant women. METHODS: A pregnant Chinese woman appeared for regular pregnancy check-ups. While monitoring blood glucose, an HbA1c test was initially performed in the pregnant Chinese woman using a CE-HPLC assay and capillary electrophoresis. Then acid gel Hb electrophoresis, a reverse dot-blot (RDB) assay, gap-PCR and DNA sequencing were applied to further evaluate the Hb variants. RESULTS: The CE-HPLC system has shown that the percentage of blood HbA1c was 5.4%, while hemoglobin electrophoresis of HbA2 and HbA0 were 1.6% and 88.7%, respectively. The capillary electrophoresis measurement results were 6.4% HbA1c, 0.4% HbA2, and 84.1% HbA0. HbA2 and HbA were 0.8% and 89.3%, respectively. Further gap-PCR, the reverse dot-blot (RDB) assay, and DNA sequencing showed that the pregnant woman had HbH-CS (αα CS/--) (1800 bp and 1300 bp) and ß/ß. CONCLUSIONS: Our study demonstrates that hemoglobin variants, such as HbH-CS can affect capillary electrophoresis results. An HbH-CS diagnosis in a pregnant woman is clinically significant. Laboratories should be cautious in using the CE-HPLC assay to evaluate HbA1c results in the presence of hemoglobin variants. Our findings highlight the strong discriminatory power of capillary electrophoresis for various Hb variants.

Detection and analysis of resistance mutations of hepatitis B virus.

OBJECTIVE: To investigate the relationships of resistance mutations of hepatitis B virus (HBV) with replication and genotypes of HBV and to understand the common resistance mutations and mutation pattern. METHODS: The mutation patterns related to resistance to nucleoside drugs were analyzed, and the relationships of resistance mutations with HBV genotypes, Ct value, HBeAg, alanine aminotransferase (ALT), age and gender were evaluated. RESULTS: Genotype B was found in 52 patients (73.2%) and genotype C in 19 patients (26.8%). In addition, 32 patients (45.07%) had resistance mutations at different loci, of which single base mutation accounted for 56.25% (18/32) and multi-base mutation for 43.75% (14/32). Of single base mutation, L180M, M204I, M204V and V173L had higher prevalence, and the incidence of L180M was closely related to the genotype of HBV. L180M, M204I and M204V were associated with the resistance to lamivudine and telbivudine; L180M, M204I, M204V and V173L were associated with the resistance to entecavir; A181T, N236T and N/H238T were related to the resistance to adefovir. Of multi-base mutations, L180M combined M204V had a high prevalence and were frequently found in patients with resistance to lamivudine and telbivudine. There was cross-resistance between lamivudine and telbivudine, between lamivudine and entecavir, and between entecavir and telbivudine. The Ct value of HBV DNA, HBeAg, ALT, age and gender were comparable among patients with different resistance mutations and HBV genotypes. CONCLUSION: Detection of mutations of multiloci resistance genes is helpful for timely identification of HBV resistance and the clinical anti-virus therapy.

A 4.6 kb genomic duplication on 20p12.2-12.3 is associated with brachydactyly type A2 in a Chinese family.

BACKGROUND: Brachydactyly type A2 (BDA2) is an autosomal dominant disorder. It was recently reported that a 5.9 kb duplication and a 5.5 kb duplication in the region 20p12.2-12.3 are associated with BDA2 in two European families. OBJECTIVE: To characterise a 6-generation Chinese family with 16 members affected by BDA2 and map the gene to 20p12.2-12.3. METHODS AND RESULTS: A 4.6 kb duplication downstream of the bone morphogenetic protein 2 (BMP2) was identified in the family. The duplication co-segregated with the phenotype and was absent in unaffected family members and control subjects. Coding and splice-site mutations of all annotated genes in the critical region were also excluded. The duplication partially overlaps with the reported duplications but has a different breakpoint. The most conserved 2.1 kb fragment in the duplication was cloned into the pGL3-promoter vector downstream of the firefly luciferase reporter gene in the 5' to 3' orientation and transfected into osteosarcoma U-2OS and Hela cells. A reduced luciferase activity was observed. CONCLUSION: The smallest duplication is described, which partially overlaps the reported duplications but has a different breakpoint, and its association with BDA2 in a Chinese family is confirmed. The results also provide evidence for cis-regulatory sequences in the duplication 3' of BMP2.