Jan-Gowth Chang

Association of CTLA4 gene A–G polymorphism with type 1 diabetes in Chinese children†

The CTLA4 (cytotoxic T lymphocyte associated antigen‐4) gene encodes the T cell receptor involved in the control of T cell proliferation and mediates T cell apoptosis. Thus it is a strong candidate gene for T cell‐mediated autoimmune disease. There is polymorphism at position 49 in exon 1 of the CTLA4 gene, providing a A–G exchange. This polymorphism is reportedly associated with type 1 diabetes in Caucasians but not in a small data set of Chinese. We wished to test this polymorphism in a larger and more homogeneous data set of Chinese children with type 1 diabetes and normal adult controls.

Polymorphism of Transmembrane Region of MICA Gene and Kawasaki Disease

Kawasaki disease is a febrile disease of children complicated with vasculitis of the coronary arteries and potential aneurysm formation. It has been recognized worldwide and appears to be increasing in frequency. Studies have found that Kawasaki disease is associated with major histocompatibility complex (MHC) class I B antigens. The MHC-class-I-chain-related gene A (MICA) is located near HLA-B. It has a triplet repeat microsatellite polymorphism in the transmembrane region. We investigated the microsatellite polymorphism in children with Kawasaki disease and controls. Seventy children (46 boys), age at diagnosis 1.68 ± 1.69 years, with Kawasaki who were treated with aspirin as well as intravenous gamma-globulin were enrolled. Control subjects consisted of 154 children (87 boys), age 2.81 ± 2.12 years. Phenotype frequency of allele A4 in patients with aneurysm formation was significantly lower than in patients without aneurysms [relative risk (RR) = 0.06, 95% confidence interval (CI) = 0.01–0.48, p = 0.00469, pc = 0.0232] and showed a similar tendency when compared with controls. Gene frequency of allele A4 was also significantly lower in patients who developed aneurysms than in patients who did not (RR = 0.07, 95% CI = 0.01–0.57, p = 0.0057, pc = 0.0282). Gene frequency of allele A5 showed a tendency to be higher in patients who developed aneurysms than in controls (RR = 2.35, 95% CI = 0.98–5.63, p = 0.0486, pc = 0.220). Allele A5.1 tended to be negatively associated with Kawasaki disease (RR = 0.57, 95% CI = 0.35–0.93, p = 0.022, pc = 0.105). Our study showed that allele A4 was negatively associated with coronary aneurysm formation in Kawasaki disease. This suggests that allele A4 protects the children with Kawasaki disease from developing coronary aneurysms after aspirin and gamma globulin therapy.

Mutation analysis of the PTEN/MMAC1 gene in cancers of the digestive tract

The 10q23.3 gene PTEN (phosphatase and Tensin homologue deleted on chromosome 10) or MMAC1 (mutated in multiple advanced cancers 1) was recently reported to undergo frequent mutation, including mutations and deletions in multiple advanced cancers. This study showed that the aberrant transcripts of this gene are frequently found in cancers of the digestive tract, paired non-cancerous tissues and normal peripheral mononuclear cells. Sequence analysis of the aberrant transcripts revealed three types of deletions: (i) a deletion junction with a splicing-like donor or acceptor sequence; (ii) several-base homology near or between the donor acceptor site at the deletion junction; and (iii) deletion with insertion. From these results, it is suggested that aberrant transcripts of PTEN/MMAC1 found by nested reverse transcription-polymerase chain reaction are a common (or natural) phenomenon unrelated to oncogenesis. The mechanism producing these aberrant transcripts needs further investigation. Using single-strand conformation polymorphism and direct sequencing to analyse for small base changes of the genomic DNA of the PTEN/MMAC1 gene revealed no point mutations or small base changes.

Polymorphism in the transmembrane region of the MICA gene and type 1 diabetes.

Although MHC class II genes have a stronger association with type 1 diabetes than MHC class I genes, studies have shown that MHC class I molecules play an independent role in the etiology of type 1 diabetes, and the existence of susceptibility genes within a segment of MHC between the HLA-B and TNF genes has been predicted, where MHC class I chain-related gene A (MICA) resides. MICA has a triplet repeat polymorphism in the transmembrane region consisting of five alleles. We analyzed this polymorphism in 162 unrelated children (82 boys) with type 1 diabetes (age at diagnosis 7.01 +/- 3.76 yr) and 154 randomly selected unrelated children (87 boys), age 2.81 +/- 2.12 yr. Phenotype frequency of allele A9 in children with type 1 diabetes was significantly higher than in controls (RR = 2.42, 95% CI = 1.52-3.85, p = 0.000162, pc = 0.00081). Gene frequency of allele A9 was also significantly higher in children with type 1 diabetes when compared with control children (RR = 2.73, 95% CI = 1.85-4.03, p = 2.62 x 10(-7), pc = 1.31 x 10(-6)). This study demonstrates that MICA allele A9 confers risk of type 1 diabetes.

The promoter region of the CTLA4 gene is associated with type 1 diabetes mellitus.

The CTLA4 (cytotoxic T lymphocyte associated antigen-4) gene encodes the T cell receptor involved in the control of T cell proliferation and mediates T cell apoptosis. C-T polymorphism is present at position -318 from the ATG start codon in the promoter region of the gene. We report a study on the polymorphism in 347 unrelated children with type 1 diabetes mellitus (DM) (age at diagnosis 7.2+/-3.8 years) and their 260 healthy siblings as controls. Genotype C/C conferred a risk of type 1 DM (RR = 2.02, 95% CI 1.32-3.10, pc = 0.0033). The gene frequency of the C allele was higher in patients (RR = 1.91, 95% CI 1.28-2.84, pc = 0.0026). The gene frequency and phenotype frequency of the T allele were negatively associated with type 1 DM (RR = 0.52, 95% CI 0.35-0.78, pc = 0.0026 and RR = 0.49, 95% CI 0.32-0.76, pc = 0.0022, respectively). The frequency of genotype C/T was lower in patients (RR = 0.50, 95% CI 0.32-0.78, pc = 0.0051). This study demonstrates that nucleotide -318 C-T polymorphism of the CTLA4 gene is associated with type 1 DM. The promoter allele -318 C confers a risk of type 1 DM but allele -318 T confers protection against this disease.

Mutation analysis of the putative tumor suppressor gene PTEN/MMAC1 in hepatocellular carcinoma.

Abstract Loss of heterozygosity of chromosome 10q has been reported in hepatoma. Areas with a high rate of loss of genetic material could harbor putative tumor suppressor genes. PTEN/MMAC1, a candidate tumor suppressor gene located at chromosome 10q23.3, has recently been identified and found to be homozygously deleted or mutated in several different types of human tumors. To determine whether the PTEN/MMAC1 gene is a target of 10q loss of heterozygosity in hepatoma, we examined 42 primary hepatomas for mutations in PTEN/MMAC1 by using nested reverse transcriptase polymerase chain reaction (RT-PCR) of the RNA and single-stranded conformation polymorphism (SSCP) analysis of all genomic exons. Although 2 of 42 hepatoma tissues had aberrant transcripts, 5 matched noncancerous liver tissues also had aberrant transcripts. Southern blot analysis of the entire genomic DNA revealed no genomic change. Therefore, like the TSG101 or FHIT gene, aberrant transcripts of PTEN/MMACl using the nested RT-PCR method were a common phenomenon for both cancerous and noncancerous liver tissues, which may not be related to oncogenesis. None of the 42 cases had small deletions, point mutations, or insertions. Our results suggest that the PTEN/MMAC1 gene may not play a role in the pathogenesis of hepatoma.

G protein β3 subunit variant and essential hypertension in Taiwan — a case–control study

Recent studies have shown that a C825T polymorphism of the gene encoding the G protein β3 subunit contributes to the genesis of essential hypertension. However, the link between the gene and blood pressure is not consistently found in different populations. The aim of the present study is to investigate this issue in Taiwan. We analyzed the allelic status in 302 hypertensive (age, 60±11 years; male/female, 136/166) and 199 normotensive subjects (62±15 years; male/female, 90/109). Our result showed that the T allelic was more frequently seen in the hypertensive group than the normotensive, but the difference did not reach statistic significance (56.5 vs. 54.3%, P>0.1). Subsequent analysis demonstrated a similar trend in the female (58.7 vs. 53.7%, P>0.1) but a reverse trend in the male (53.7 vs. 55%, P>0.1). Another finding was that the T allele frequency in all the groups was over 50%, markedly higher than those reported in whites. In conclusion, the observation suggests that the polymorphism in the G protein gene is not likely to play an important role in the manifestation of high blood pressure in Taiwan.

Mutation analysis of the putative tumor suppression gene PTEN/MMAC1 in sporadic breast cancer

PTEN/MMAC1, a potential human tumor suppressor gene, has been found to have inactivating mutations in several types of cancer, including breast cancer. The incidence of breast cancer in Chinese is quite low in comparison with Caucasians, and genetic factors may play some roles. To further determine the role of PTEN/MMAC1 in breast cancer in Chinese, we used loss of heterozygosity (LOH), single strand conformation polymorphism (SSCP) with direct sequencing of variant bands, and Southern blot analysis methods to analyze mutations in PTEN/MMAC1 in 52 cases of breast cancer. None had LOH at chromosome 10q23.3. One mutation was identified, a somatic 3‐base deletion, in one case. Our results suggest PTEN/MMAC1 does not play a major role in the development of sporadic breast cancer.

Aberrant TSG101 transcripts in acute myeloid leukaemia

Recently, a tumour susceptibility gene, TSG101, has been identified at chromosome 11p15. A large intragenic deletion of this gene has been demonstrated in primary breast tumours. To evaluate the role of the TSG101 gene in leukaemia, bone marrow and/or peripheral blood from 68 acute myeloid leukaemia patients, five haemopoietic cell lines (HL60, U937, Raji, KG‐1, K562) and 30 normal controls were analysed by reverse transcription of the TSG101 mRNA, followed by PCR amplification and sequencing of the products. The results showed aberrant TSG101 transcripts in 24/68 (35%) acute myeloid leukaemia (AML) patients, all of the cell lines (100%) and 3/30 (10%) normal controls. Our study indicated that the abnormal transcripts may have resulted from aberrant RNA splicing as evidenced by these aberrant transcripts. Also, normal full‐length transcripts were present in all specimens examined. The aberrant transcript occurred more frequently in the AML and cell lines. However, because aberrant transcripts of TSG101 were also found in the normal controls, the role of TSG101 as a tumour suppressor gene should be evaluated carefully.

Prenatal prediction of spinal muscular atrophy in Chinese

We used linkage analysis, non‐isotope SSCP (single‐strand conformation polymorphism) and PCR‐RFLP (polymerase chain reaction‐restriction fragment length polymorphism) for prenatal diagnosis of spinal muscular atrophy (SMA). A total of 26 cases from 20 SMA families (16, type 1 and 4) were evaluated. 5 out of 26 fetuses were affected and, following genetic counselling, the parents decided to terminate the pregnancies. Aborted fetal tissues were examined and the diagnosis was confirmed in each case. The 21 unaffected cases were either normals (12 cases) or carriers (9 cases). These children have been followed for six months to two and a half years. No false‐negative or false‐positive results on prenatal testing were found. We conclude that prenatal diagnosis of SMA is reliable and accurate. Copyright