Relationship between reduced folate carrier gene polymorphism and non-syndromic cleft lip and palate in Indian population

Objective: Folate metabolism involves absorption, transport, modifications and interconversions of folates. The reduced folate carrier does not participate directly in folate metabolism but plays a major role in intracellular transport of metabolically active 5-methyltetrahydrofolate and maintains the intracellular concentrations of folate. The purpose of this study was to identify the prevalence of reduced folate carrier 1 (RFC1) A80G polymorphism and to further delineate its association with non-syndromic cleft lip and palate (NSCLP) in a south Indian population.

Methods: In the present case-control study, we studied RFC1 gene A80G polymorphism to evaluate its impact on NSCLP risk in south Indian population. Blood samples of 142 cases with NSCLP and 141 controls were collected and genotyped using PCR-RFLP.

Results: The genotype distribution in the control group followed Hardy–Weinberg equilibrium (p?=?0.633). The G allele frequency of cases was 64.8% (184/284) and was significantly lower than that found in the control group 56.4% (160/282). The genotype distributions between NSCLP cases and controls was not significantly different (p?=?0.131). The allelic model significantly increased the risk of NSCLP (G versus A; OR?=?1.40; 95% CI: 1.00–1.97; p?=?0.050). In subgroup analysis, the A80G variant showed significant association for the CLP group in dominant and allelic models.

Conclusions: Altogether, our findings support the hypothesis that RFC1 A80G variant may contribute to NSCLP susceptibility in a south Indian population.     

When and How Should We Revascularize Patients With Atherosclerotic Renal Artery Stenosis?

Atherosclerotic renal artery stenosis is the leading cause of secondary hypertension and may lead to resistant (refractory) hypertension, progressive decline in renal function, and cardiac destabilization syndromes (pulmonary edema, recurrent heart failure, or acute coronary syndromes) despite guideline-directed medical therapy. Although randomized controlled trials comparing medical therapy with medical therapy and renal artery stenting have failed to show a benefit for renal artery stenting, according to comparative effectiveness reviews by the Agency for Healthcare Research and Quality, the trials may not have enrolled patients with the most severe atherosclerotic renal artery stenosis, who would be more likely to benefit from renal stenting. Because of limitations of conventional angiography, it is critical that the hemodynamic severity of moderately severe (50% to 70%) atherosclerotic renal artery stenosis lesions be confirmed on hemodynamic measurement. The authors review techniques to optimize patient selection, to minimize procedural complications, and to facilitate durable patency of renal stenting. The authors also review the current American College of Cardiology and American Heart Association guidelines and the Society for Cardiovascular Angiography and Interventions appropriate use criteria as they relate to renal stenting.     

Genotyping of the reduced folate carrier‐1 c.80G>A polymorphism by pyrosequencing technology: Importance of PCR and pre‐PCR optimization

When developing a genotyping assay by Pyrosequencing^? technology for the RFC1 (SLC19A1) c.80G>A polymorphism (rs1051266), unequal peak heights in the pyrograms were observed, probably due to unequal amplification of the mutated and wild‐type alleles. This rarely occurring problem could potentially render assignment of heterozygous genotypes uncertain. When the PCR conditions were studied, it was found that substitution of the dGTP nucleotide in the master mix by dGTP and dITP in proportion 1:1 largely overcame this problem. Heat denaturation of the DNA at 95°C before PCR also counteracted the problem. A combination of these two modifications of the standard pyrosequencing PCR protocol gave the best results. We conclude that, with these modifications, the RFC1 c.80G>A SNP can be reliably assayed by pyrosequencing.     

Further studies on the interaction of nonpolyglutamatable aminopterin analogs with dihydrofolate reductase and the reduced folate carrier as determinants of in vitro antitumor activity

Thirteen structural analogs of the potent nonpolyglutamatable dihydrofolate reductase inhibitor N(alpha)-(4-amino-4-deoxypteroyl)-N(delta)-hemiphthaloyl-L-ornithine (PT523) with modifications in the side chain, the para-aminobenzoyl moiety, or the 9,10-bridge were evaluated for the ability to inhibit human recombinant dihydrofolate reductase (DHFR), to utilize the reduced folate carrier (RFC) for influx, and to inhibit the growth of CCRF-CEM human leukemia cells in culture. In spectrophotometric assays of the kinetics of the reduction of dihydrofolate by DHFR in the presence of NADPH, these compounds had K(i) values ranging from 0.2 to 1.3pM, and thus were not greatly different in potency from the parent drug PT523. By comparison, the K(i) values of aminopterin (AMT), methotrexate (MTX), and 10-ethyl-10-deazaaminopterin (EDX) were 3.7, 4.8, and 11pM. In assays of competitive inhibition of [3H]MTX influx into CCRF-CEM cells, the K(i) values ranged from 0.21 to 7.3 micro M, as compared with 0.71, 5.4, and 1.1 micro M for PT523, AMT, and EDX. The K(t) for MTX was also re-analyzed and found to be 4.7 micro M, in better agreement with the literature than our previously reported value of 7.1 micro M. The IC(50) values of these compounds as inhibitors of the growth of CCRF-CEM cells after 72hr of drug exposure ranged from 0.53 to 55nM, and were qualitatively consistent with the other results.     

Decreased expression of the reduced folate carrier and folypolyglutamate synthetase is the basis for acquired resistance to the pemetrexed antifolate (LY231514) in an L1210 murine leukemia cell line

Pemetrexed (LY231514) is a new-generation antifolate that, in its polyglutamyl forms, is a potent inhibitor of thymidylate synthase and glycinamide ribonucleotide formyltransferase (GAR transformylase). This study explored the mechanisms of resistance to pemetrexed in L1210 murine leukemia cells using chemical mutagenesis with 5-formyltetrahydrofolate (5-formylTHF) as the growth substrate. A cell line, MTA-13, was identified that was 8.5-fold resistant to pemetrexed with comparable cross-resistance to ZD1694 (Tomudex) and lesser cross-resistance (5-fold) to ZD9331 [(2S)-2-(O-fluoro-p-[N-(2,7-dimethyl-4-oxo-3,4-dihydro-quinazolin-6-ylmethyl)-N-(prop-2-ynyl)amino]benzamido)-4-(tetrazol-5-yl)-butyric acid], DDATHF (dideazatetrahydrofolate) (3.5-fold), and methotrexate (MTX) (2.7-fold) but comparable sensitivity to trimetrexate. Influx of pemetrexed, MTX, and 5-formylTHF into MTA-13 cells was decreased by 56, 47, and 38% compared to wild-type cells. Folate receptor expression was negligible in both cell lines. Net drug uptake declined within 15min to a slower, constant rate over the next 45min, reflecting the rate of accumulation of pemetrexed polyglutamate derivatives. This rate in the MTA-13 line was half that of the wild-type cells. Accumulation of 50nM [3H]pemetrexed, 25nM [3H]5-formylTHF, or 50nM [3H]DDATHF after 3 days was decreased to 35, 46, and 56% the level of L1210 cells. The reduced folate carrier (RFC) message and protein were decreased by 50%, and folypolyglutamate synthetase (FPGS) message was decreased by 65% in MTA-13 cells. No mutations were detected in either protein by DNA sequence analysis. There was a slight decrease (approximately 25%) in thymidylate synthase mRNA, without mutations in the protein, and there was no change in GAR transformylase message. The data indicate that resistance to pemetrexed in the MTA-13 cell line was due to changes in both RFC and FPGS expression, two proteins that act in tandem to regulate polyglutamation of folates and antifolates in cells, resulting in cellular depletion of these active pemetrexed congeners.     

The Reduced Folate Carrier (SLC19A1) c.80G>A Polymorphism is Associated with Red Cell Folate Concentrations Among Women

Low folate status may be a consequence of suboptimal intake, transport or cellular utilization of folate and, together with elevated homocysteine, is a recognized risk factor or marker for several human pathologies. As folate transport across cell membranes is mediated in part by the reduced folate carrier (RFC1), variants within SLC19A1, the gene that encodes RFC1, may influence disease risk via an effect on folate and/or homocysteine levels. The present study was undertaken to assess the association between the SLC19A1 c.80G>A polymorphism and folate/homocysteine concentrations in healthy young adults from Northern Ireland.
The SLC19A1 c.80G>A polymorphism was not strongly associated with either serum folate or homocysteine concentrations in either men or women. However, in women, but not in men, this polymorphism explained 5% of the variation in red blood cell (RBC) folate levels ( P = 0.02). Relative to women with the SLC19A1 c.80GG genotype, women with the GA and AA genotypes had higher RBC folate concentrations. Consequently, compared to women with the SLC19A1 c.80GA and AA genotypes, women who are homozygous for the 80G allele may be at increased risk of having a child affected with a neural tube defect and of developing pathologies that have been associated with folate insufficiency, such as cardiovascular disease.     

非专用线圈在MRI乳腺检查中的应用研究

目的:探讨非专用线圈在乳腺MRI检查中的应用价值。方法:运用乳腺专用线圈、SPINE matrix线圈两种检查方法扫描乳腺健康志愿者126例,Wilcoxon检验对126例健康志愿者扫描所得两组图像质量进行比较分析;利用t检验分析两组图像SNR;采用卡方检验分析两组横轴位扫描图像范围(腋窝淋巴组织)。结果:运用乳腺专用线圈和SPINEmatrix线圈两种检查方法图像质量、SNR比较差异无显著意义,SPINE matrix线圈横轴位显示双侧腋窝淋巴组织优于专用线圈,差异有显著意义。结论:SPINE matrix线圈可以达到等同乳腺专用线圈的应用效果,同时可以较好显示双侧腋窝组织。     

Reduced folate carrier gene G80A polymorphism is associated with an increased risk of gastroesophageal cancers in a Chinese population

Low folate intake has been associated with an increased risk of both oesophageal and gastric cancers. Reduced folate carrier (RFC1, also named SLC19A1) is an essential folate transporter and functions as a bidirectional anion exchanger, taking up folate cofactors and exporting various organic anions. A G80A polymorphism in RFC1 gene has been shown to be associated with alterations in folate and homocysteine metabolism in healthy individuals. In this study, we hypothesised that genetic variants in RFC1 may modulate risk of oesophageal cancer (EC) and gastric cancer (GC). To test this hypothesis, we evaluated the associations of the G80A polymorphism of RFC1 with EC and GC risk in a case-control study of 216 EC and 633 GC cases and 673 cancer-free controls in a Chinese population. We found that compared with the 80GG/GA genotypes in the recessive model, the variant homozygote RFC1 80AA was associated with a significantly increased risk of EC (adjusted odds ratio (OR) = 1.80, 95% confidence interval (CI) = 1.29–2.51), GC (adjusted OR = 1.59, 95% CI = 1.25–2.02) and EC and GC combined (adjusted OR = 1.63, 95% CI = 1.30–2.04). In the dominant model, the risk associated with RFC1 80AA was also elevated in EC (OR = 1.35, 95% CI = 0.91–1.99), GC (OR = 1.43, 95% CI = 1.07–1.91) and EC and GC combined (adjusted OR = 1.40, 95% CI = 1.07–1.83), compared with the 80GG genotype. The stratification analyses showed that effects of the RFC1 80AA genotype were more evident in subgroups of relatively older (60 years), female, non-smokers, and non-drinkers both in EC and GC. Although the exact biological mechanism of this association remains to be explored, our findings suggest possible involvement of RFC1 variant in the susceptibility of EC and GC. Further large and functional studies are needed to confirm our findings.     

Synthesis of the Gln1-facteur thymique serique and its evaluation with human E-rosette bioassay

Synthesis of an analog of facteur thymique serique (FTS) containing a glutaminyl residue in position 1 of the peptide chain is described. Like the original FTS, Gln¹-FTS is able to induce T cell markers on lymphocytes in vitro in a picogram dosage range. The human E-rosette bioassay was adopted for the evaluation of biological activity of Gln¹-FTS. It was also shown that the latter method can be used to evaluate the presence of target cells for FTS activity in peripheral blood lymphocytes in humans.