Increase in intracellular Zn concentration by thimerosal in rat thymocytes: Intracellular Zn release induced by oxidative stress

Thimerosal (TMR), an ethylmercury-containing preservative in pharmaceutical products, was recently reported to increase intracellular Zn²⁺ concentration. Therefore, some health concerns about the toxicity of TMR remain because of physiological and pathological roles of Zn²⁺. To reveal the property of TMR-induced increase in intracellular Zn²⁺ concentration, the effect of TMR on FluoZin-3 fluorescence, an indicator of intracellular Zn²⁺, of rat thymocytes was examined. TMR at concentrations ranging from 0.3 μM to 10 μM increased the intensity of FluoZin-3 fluorescence in a concentration-dependent manner under external Ca²⁺- and Zn²⁺-free condition. The threshold concentration was 0.3–1 μM. The increase in the intensity was significant when TMR concentration was 1 μM or more. N,N,N′,N′-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), a chelator for intracellular Zn²⁺, completely attenuated the TMR-induced augmentation of FluoZin-3 fluorescence. Hydrogen peroxide (H₂O₂) and N-ethylmaleimide, reducing cellular thiol content, significantly increased FluoZin-3 fluorescence intensity and decreased 5-chloromethylfluorescein (5-CMF) fluorescence intensity, an indicator for cellular thiol. The correlation coefficient between TMR-induced augmentation of FluoZin-3 fluorescence and attenuation of 5-CMF fluorescence was −0.882. TMR also attenuated the 5-CMF fluorescence in the presence of TPEN. Simultaneous application of H₂O₂ and TMR synergistically augmented the FluoZin-3 fluorescence. It is suggested that TMR increases intracellular Zn²⁺ concentration via decreasing cellular thiol content.     

Pharmacogenomics of human ABC transporters: detection of clinically important SNPs by SmartAmp2 method

Genetic polymorphisms and mutations in drug metabolizing enzymes, transporters, receptors, and other drug targets (e.g., toxicity targets) are linked to inter-individual differences in the efficacy and toxicity of many medications as well as risk of genetic diseases. Validation of clinically important genetic polymorphisms and the development of new technologies to rapidly detect clinically important variants are critical issues for advancing personalized medicine. A key requirement for the advancing personalized medicine resides in the ability of rapidly and conveniently testing patients' genetic polymorphisms and/or mutations. We have recently developed a rapid and cost-effective method, named Smart Amplification Process 2 (SmartAmp2), which enables us to detect genetic polymorphisms or mutations in target genes within 30 to 45 min under isothermal conditions without DNA isolation and PCR amplification. Detection of mutations or single nucleotide polymorphisms (SNPs) in human ABC transporter genes is becoming more important, since their functional impairments are reportedly associated with inherited diseases. Thus, certain genetic polymorphisms of ABC transporters are considered important biomarkers for diagnosis of inherited diseases and/or risk of drug-induced adverse reactions. In this review article, we will present the new technology of the SmartAmp2 method and its clinical applications for detection of SNPs in human ABC transporter genes, i.e., ABCC4 and ABCC11.     

Eccrine poromatosis associated with polychemotherapy

Eccrine poroma frequently occurs as a solitary tumour, and only a few reports have described the occurrence of multiple lesions. Multiple eccrine poromas, or eccrine poromatosis, may occur in patients who have undergone radiotherapy and/or polychemotherapy. We report here four cases of multiple eccrine poromas in patients who were either undergoing, or had undergone, intensive chemotherapy (from 6 months to 16 years prior to onset). Three patients had non-Hodgkin's lymphoma and one had malignant fibrous histiocytosis. The number of lesions varied from 3 to >?20 in each patient, and all the lesions occurred on non-irradiated skin. The histopatho-logical features were consistent with those of eccrine poroma, Pinkus type. In addition to radiation therapy, intensive chemotherapy may play a role in the pathogenesis of multiple eccrine poromas even many years after treatment.     

PPARγ激动剂对T细胞增殖及破骨细胞形成的影响

目的:观察过氧化物酶体增殖物激活受体γ(peroxisome proliferator activated receptor-γ,PPARγ)激动剂15d-PGJ2对小鼠T细胞增殖、T细胞分泌的细胞因子表达及破骨样细胞形成的影响,并探讨其可能的作用机制。方法:体外分离伴放线放线杆菌(A.actinomycetetemcomitans,Aa)免疫的BALB/c小鼠颈部淋巴细胞,提取T细胞进行体外扩增,分别加入浓度为0、1×10-8、1×10-7,1×10-6,1×10-5mol/L的15d-PGJ2进行干预。培养3d后,3H-Tdr掺入法测定T细胞的增殖反应;ELISA法测定细胞上清中核因子-κB受体活化因子配体(receptor activator of NF-κB ligand,RANKL)、TNF-α和IL-10的表达水平;取扩增的T细胞上清与RAW 264.7细胞共同培养后,抗酒石酸酸性磷酸酶染色(tartrate resistant acid phosphatase,TRAP)测定破骨样细胞的形成。采用SPSS 11.0软件包对数据进行统计学分析。结果:以1×10-5mol/L 15d-PGJ2处理的小鼠T细胞3d后,与对照组相比,T细胞增殖显著受抑;上清中RANKL、TNF-α的表达水平下降,IL-10无显著变化;TRAP染色阳性细胞数减少,具有统计学意义(P<0.05),且呈浓度依赖性。结论:PPARγ激动剂15d-PGJ2可抑制T细胞增殖,减少炎性因子分泌,降低破骨样细胞的形成,提示PPARγ配体在抑制T细胞诱导的骨吸收方面发挥积极作用。     

A comparative study of clinical and radiological outcomes of dorsally angulated, unstable distal radius fractures in elderly patients: intrafocal pinning versus volar locking plating

PURPOSE: To compare the clinical and radiological outcomes of intrafocal pinning (IFP) and volar locking plating (VLP) of dorsally angulated, unstable distal radius fractures in elderly patients. METHODS: The subjects were 62 consecutive patients over 60 years of age with dorsally angulated, unstable distal radius fractures treated with IFP or VLP. Bone mineral density (BMD) of the lumbar spine was measured by dual-energy x-ray absorptiometry at first examination. The range of motion and grip strength were measured at follow-up examinations, and ulnar variance (UV) was measured on radiographs at baseline and follow-up postoperative examinations. RESULTS: There were no notable differences in gender, age, follow-up period, baseline UV, BMD, and AO classification between IFP and VLP groups. There was no difference between the UV in VLP immediately after surgery and at the final follow-up examination; however, IFP showed a significant loss of reduction as measured by UV. In patients with UV more than 5 mm or BMD less than 70% of young adult mean (YAM) at first examination, UV increased again at the final follow-up examination in IFP, while surgically corrected UV was maintained in VLP, independent of the degree of baseline UV and BMD. VLP resulted in earlier recovery of postoperative range of motion and grip strength compared with IFP. The range of flexion and the grip strength value were significantly larger in VLP at final examination. CONCLUSIONS: VLP, but not IFP, can maintain surgically corrected UV in distal radius fractures, independent of the degrees of initial UV and BMD. VLP enhances earlier recovery in range of motion and grip strength than IFP. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic II.     

Effects of beta-blocker therapy on high sensitivity c-reactive protein, oxidative stress, and cardiac function in patients with congestive heart failure

BackgroundIt is uncertain whether β-blocker therapy affects serum C-reactive protein (CRP) level in patients with congestive heart failure (CHF). We attempted to determine if β-blocker therapy decreases serum CRP production and to correlate the production with biomarkers and cardiac function in such patients.Methods and ResultsFifty-two patients with mild to moderate CHF with a left ventricular ejection fraction (EF) <40% were enrolled. They were randomly assigned to metoprolol or carvedilol treatment groups. The CRP concentration decreased significantly in patients with higher baseline CRP concentration, but not in those with lower baseline CRP concentrations. There was an inverse correlation between ΔCRP and ΔEF 16 weeks after the start of β-blocker therapy for patients with higher baseline CRP concentrations. In patients with higher baseline concentrations, CRP decreased in patients who received carvedilol, but not in those who received metoprolol. Plasma lipid peroxide (LPO) concentration significantly decreased, and there was an inverse correlation between ΔCRP and ΔLPO 16 weeks after the start of therapy.ConclusionsAdministration of β-blockers is associated with attenuation of inflammatory marker in certain patients with CHF. The antioxidant effects of β-blockers, especially carvedilol, may play a role in mediating the phenomenon.     

Correlation between parathyroid hormone, bone alkaline phosphatase and N-telopeptide of type 1 collagen in diabetic and non-diabetic haemodialysis patients

BACKGROUND: Haemodialysis (HD) patients with diabetes mellitus often have renal osteodystrophy (ROD) characterized by reduced bone turnover, but little is known about the correlation between bone formation and bone resorption in this population. METHODS: The authors measured serum parathyroid hormone (PTH), bone alkaline phosphatase (BAP), N-telopeptides of type 1 collagen (NTx) and fasting glucose in 48 patients with diabetic nephropathy (DN) and 80 patients with glomerulonephritis (non-DN) who had received or=5 years HD (r = 0.568) this correlation was similar to that in the non-DN group (r = 0.653), whereas there was no significant correlation in those receiving <5 years HD. Patients receiving >or=5 years HD had a comparable glucose level (111.1 +/- 19.2 mg/dL) to the non-DN group, whereas those receiving <5 years had a higher level (196.1 +/- 53.1 mg/dL). CONCLUSION: Differences in the interaction between bone cells between DN and non-DN patients are one potential cause of lower bone turnover in the former group. Research of this correlation is needed to increase understanding of the complexities of bone metabolism in DN patients.     

Diffuse large B-cell lymphoma in the thyroid gland associated with primary hyperparathyroidism

We report a case of primary hyperparathyroidism associated with a malignant lymphoma in the thyroid gland. A 68-year-old woman was admitted to hospital with a cervical mass. Ultrasound and computed tomography (CT) revealed a hypoechoic, multinodular tumor in the left thyroid gland. A gallium-67 citrate scintigram revealed intense radioisotope uptake in the thyroid tumor. Histological examination of biopsy specimens indicated that this tumor was a large B-cell lymphoma. The coexistence of parathyroid adenoma in this patient was revealed by a sestamibi scintigram, performed prior to chemotherapy. Following the complete remission of the lymphoma by chemotherapy, we carried out an excision of the single parathyroid adenoma. To our knowledge, this is the first report to describe a malignant thyroid lymphoma associated with primary hyperparathyroidism.     

Re-evaluation and functional classification of non-synonymous single nucleotide polymorphisms of the human ATP-binding cassette transporter ABCG2

Impacts of genetic polymorphisms of the ATP-binding cassette (ABC) transporter BCRP/MXR1/ABCP (ABCG2) on drug response have been implicated; however, the hitherto reported data involve some inconsistencies. To re-evaluate the effect of single nucleotide polymorphisms (SNP) of ABCG2 in vitro, we created a total of seven variant cDNAs (V12M, Q141K, F208S, S248P, F431L, S441N and F489L) by site-directed mutagenesis and stably expressed each of them in Flp-In-293 cells using the Flp recombinase system. Multicolor fluorescence in situ hybridization mapping analysis revealed that one single copy of ABCG2 cDNA was incorporated into the telomeric region of chromosome 12p. It was proven that mRNAs of those integrated ABCG2 variants were expressed evenly in Flp-In-293 cells. However, the protein expression levels varied among those variants. In particular, expression of the F208S and S441N variants was markedly low, suggesting the instability of these variant proteins. Drug resistance profiles of Flp-In-293 cells expressing two major SNP variants (V12M and Q141K) toward the drug SN-38 demonstrated that the IC50 value (drug concentrations producing a 50% reduction of cell growth) for Q141K was approximately 50% of that for wild type. The contributions of the minor SNP variants (F208S, S248P, F431L, S441N and F489L) to drug resistance toward SN-38, mitoxantrone, doxorubicin, daunorubicin or etoposide were significantly lower than wild type. Based on our functional validation, the above-mentioned non-synonymous polymorphisms as well as acquired mutants (R482G and R482T) of ABCG2 were classified into four groups. Furthermore, new camptothecin analogs synthesized by our research group had potent effects in circumventing ABCG2-mediated drug resistance without any influence from major non-synonymous polymorphisms.