Nancy M. Wang

A mouse model for spinal muscular atrophy

The survival motor neuron gene is present in humans in a telomeric copy, SMN1, and several centromeric copies, SMN2. Homozygous mutation of SMN1 is associated with proximal spinal muscular atrophy (SMA), a severe motor neuron disease characterized by early childhood onset of progressive muscle weakness. To understand the functional role of SMN1 in SMA, we produced mouse lines deficient for mouse Smn and transgenic mouse lines that expressed human SMN2. Smn−/− mice died during the peri-implantation stage. In contrast, transgenic mice harbouring SMN2 in the Smn−/− background showed pathological changes in the spinal cord and skeletal muscles similar to those of SMA patients. The severity of the pathological changes in these mice correlated with the amount of SMN protein that contained the region encoded by exon 7. Our results demonstrate that SMN2 can partially compensate for lack of SMN1. The variable phenotypes of Smn−/−SMN2 mice reflect those seen in SMA patients, providing a mouse model for this disease.

Treatment of spinal muscular atrophy by sodium butyrate.

Spinal muscular atrophy (SMA) is an autosomal recessive disease characterized by degeneration of the anterior horn cells of the spinal cord, leading to muscular paralysis with muscular atrophy. No effective treatment of this disorder is presently available. Studies of the correlation between disease severity and the amount of survival motor neuron (SMN) protein have shown an inverse relationship. We report that sodium butyrate effectively increases the amount of exon 7-containing SMN protein in SMA lymphoid cell lines by changing the alternative splicing pattern of exon 7 in the SMN2 gene. In vivo, sodium butyrate treatment of SMA-like mice resulted in increased expression of SMN protein in motor neurons of the spinal cord and resulted in significant improvement of SMA clinical symptoms. Oral administration of sodium butyrate to intercrosses of heterozygous pregnant knockout-transgenic SMA-like mice decreased the birth rate of severe types of SMA-like mice, and SMA symptoms were ameliorated for all three types of SMA-like mice. These results suggest that sodium butyrate may be an effective drug for the treatment of human SMA patients.

No evidence of correlation between mutation at codon 531 of src and the risk of colon cancer in Chinese

The protein tyrosine kinase activity of c-src proto-oncogene product, pp60(c-src), is elevated in a number of human cancers, including colon cancer. Phosphorylation of human pp60(c-src) carboxy-terminal tyrosine 530 suppresses its kinase activity. A recent report suggested that the risk of colon cancer is higher for those who carry a C-->T transition mutation on codon 531 (Gln-531-->Amber-531) of src gene. This mutation caused a prematured translation termination and up-regulated the kinase activity. To examine whether this mutation could be a risk factor for colon carcinoma in the Chinese population, we used the same PCR-based assay to analyze src genotypes of 131 colon cancers and other various types of carcinoma. No mutation was detected in all specimens that were screened in this study. Thus, mutation at Gln-531 of src gene does not seem to be involved in the development of colon cancer in Chinese ethnicity.

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.

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

Mutation Analysis of RAS Oncogenes in Oral Squamous Cell Carcinoma

Background. The mutations of RAS oncogenes have been found in a number of cancers and play an important role in oncogenesis. However, only a few studies have analyzed the mutations of N-,H-, and K- RAS in oral squamous cell carcinoma. Methods. Hotspot mutations of N-, H- and K- RAS oncogenes were analyzed by the amplified created restriction site method (ACRS), polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) and direct sequencing analysis. Results. Out of the 20 squamous cell carcinoma (SCC) specimens we studied, four had a GGC to AGC change at codon 12, which corresponds to an amino acid glycine to serine mutation of the H-RAS oncogene. No mutation was found in K-or N-RAS oncogenes. Conclusions. Our results differed from Kuo et al, which may be due to different chemical components in betel quid or to different patient populations studied. Further analysis is needed.

Mutation analysis of the Bcl 10 gene in hepatocellular carcinoma.

The Bcl 10 gene was recently discovered to be involved in the pathogenesis of lymphoma of mucosa-associated lymphoid tissue and several types of tumorous cell lines. We examined the mutation of Bcl 10 gene in 31 hepatocellular carcinomas along with their corresponding non-tumorous tissues by single strand conformation polymorphism (SSCP) and direct sequencing. The results showed that 11.3% chromosomes had codon 5 GCA to TCA mutation, 4.8% chromosomes had codon 8 CTC to CTG mutation, and 12.9% chromosomes had codon 213 GGA to GAA mutation. These mutations were found not only in the hepatoma tissues but also in paired non-cancerous tissues and the normal population. We suggest that these three changes are polymorphisms, and there is no relationship with the development of hepatocellular carcinoma.

Comparison of Different Sources of Infrared Irradiation on COS-1 Fibroblast Cell Growth

Objectives: The purpose of this study was to investigate the effects of ceramic infrared (CIR) and traditional infrared (TIR) irradiation on COS-l fibroblast cell growth and to distinguish the non-thermal effect from the thermal effect of infrared irradiation. Methods: COS-1 fibroblast cells were irradiated in a 23OC temperature controlled dark room. The cells were divided into a CIR group, a TIR group and an ambient (control) group. The cells were then irradiated at various distances from the radiation source and for various time durations. The distance of non-thermal irradiation was identified. We then counted the number of cells 24 hours after irradiation. The average cell proliferation in the three dishes was taken. Differences between cell proliferation in the CIR group and those in the TIR were compared. Results: TIR irradiation elevated the temperature much higher than CIR, especially at short distances. TIR destroyed cells at short distances because of high temperature irradiation. CIR irradiation had a more stimulating effect on fibroblast proliferation than TIR. Both non-thermal and thermal irradiation stimulated cell proliferation, but adequate heat and non-thermal irradiation provided by CIR produced optimal effects on cell growth. Conclusions: These findings indicate that duration and distance of irradiation are important factors for stimulating COS-l fibroblast cell growth. Furthermore, the non-thermal effect of CIR, which is less harmful than TIR, stimulates fibroblasts more effectively than TIR.

Mutational Analysis of the Melanocortin-4 Receptor Gene in Chinese Children with Obesity

Objectives. The melanocortin system is involved in physiological regulation of homeostasis. The melanocortin-4 receptor (MC4R) in the hypothalamus is thought to be important in controlling feeding behavior. Genetic studies have identified two mutations, Ser-30-Phe and Asp298-Asn, of MC4-R which are associated with dominant forms of inherited obesity. Methods. In order to examine whether these two mutations are associated, with the development of early-onset obesity in the Chinese population, we used a polymerase chain reaction-base analysis to analyze the MC4R genotypes of 92 obese and 185 non-obese children. Results. No mutations were detected in any of the individuals that were screened in this study. However, two novel polymorphisms, Ile-1O3 and Ile-297, were identified in both obese and normal individuals which were associated with the phenotypes under study. Conclusions. It seems that the Ser-30-Phe and Asp-298-Asn mutations do not contribute to inherited susceptibility to early-onset obesity in Chinese children.

The Relationship Between Early-onset Childhood Obesity and POMC and UCP3 Gene Variance

Background: Human Uncoupling Protein 3 (UCP3) is a new candidate gene for human obesity. UCPJ has been shown to be regulated by thyroid hormone, 3-adrenergic agonists, leptin, and fat feeding in rodents. Mutations in exon 3 (V1O2I) and exon 4 (R143X) were identified in obese and diabetic pro bands. Pro-opiomeleancortin (POMC), another obese gene, plays a central role in α-MSH regulation of food intake by activating melanocortin-4-receptors in the brain. The POMCgene codes for the prohormone pro-opiomelanocortin, the hormone suspected of playing a role in appetite and body weight regulation. Methods: We selected 100 obese and 300 non-obese children for analysis. Obesity was defined as a body mass index greater than or equal to the 95percentile. To examine whether mutations in exon 3 (V1O2I) and 4 (R143X) of the uncoupling protein gene might be factors for obesity in Taiwanese children, the target DNA fragments were amplified by polymerase chain reaction, and the genotypes were defined by restriction fragment length polymorphism. Results: No significant differences in the frequency of these mutations were found between obese and non-obese children. Conclusions: The results indicate that those mutations of the UCP3 and POMC genes do not playa role in the development of obesity in Taiwans children.