Outcomes of multitarget therapy using mycophenolate mofetil and tacrolimus for refractory or relapsing lupus nephritis.

Objectives Outcomes of systemic lupus erythematosus (SLE) have significantly improved over the years. However, when there is major organ involvement, the outcomes can still be unfavorable. Outcomes of multitarget therapy using mycophenolate mofetil (MMF) and tacrolimus in patients with SLE who were refractory to standard therapy were assessed. Methods We retrospectively reviewed the Hanyang BAE lupus cohort to identify patients with biopsy-confirmed lupus nephritis (classes III, IV, or V) who failed to either achieve complete response with standard induction therapy or those who had a lupus flare after achieving a complete response with conventional induction therapy and subsequently were switched to multitarget combination therapy with MMF and tacrolimus. Outcomes, including renal response, proteinuria, glomerular filtration rate, serum albumin, anti-dsDNA antibody level, Systemic Lupus Erythematosus Disease Activity Index (SLEDAI), and complements, were assessed at six and 12 months. Results Twenty-nine patients, including 12 who failed to achieve a complete response at 12 months after initial conventional induction therapy and 17 with lupus flare after achieving a complete response at 12 months and treated with multitarget therapy, were included in the analysis. At six months, 53.9% of the patients showed a response, with 15.4% of patients showing a complete response and 38.5% of patients showing a partial response. At 12 months, 55.5% of patients exhibited a response (with complete and partial response in 25.9% and 29.6%, respectively). The dosage of steroids was significantly decreased at six months compared with baseline and was maintained at 12 months. Proteinuria, anti-double-stranded DNA antibody positivity, as well as C3 and C4 levels improved after treatment and persisted until 12 months, but were not significant. SLEDAI also improved. Outcomes were significantly better in patients who had a complete response but later had a flare, resulting in the use of multitarget therapy and achieving a subsequent complete response. Conclusions Multitarget therapy with MMF and tacrolimus can be a reasonable option in refractory lupus nephritis patients who failed to show adequate response to conventional induction therapy or who had flares during maintenance therapy. This treatment can help patients achieve a renal response and reduce the use of steroids.

Lupus nephritis is associated with more corticosteroid-associated organ damage but less corticosteroid non-associated organ damage.

Objective The objective of this study was to investigate the association of lupus nephritis on organ damage and mortality in patients with systemic lupus erythematosus (SLE). Methods A total of 1112 patients with SLE were investigated. Lupus nephritis was defined as a proteinuria based on the 1997 American College of Rheumatology criteria. Damage was assessed using the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index. The associations of lupus nephritis with overall, non-renal, corticosteroid-associated, and non-associated damage were analyzed using logistic regression. The age-adjusted and sex-adjusted standardized mortality ratio was evaluated in patients with and without lupus nephritis. Results The prevalence of lupus nephritis in patients with SLE was 46.3%. Patients with lupus nephritis had a higher percentage of overall cumulative damage than patients without lupus nephritis (51.5% vs. 35.7%, p < 0.001). The odds ratio was 1.40 after adjusting for age at SLE diagnosis, sex, disease duration, anti-malarial agents, immunosuppressive agents and cumulative corticosteroid dose. Among non-renal damage, the odds of corticosteroid-associated damage were higher (2.06, 95% confidence interval (CI) 1.43-2.96) whereas the odds of non-associated damage were lower (0.50, 95% CI 0.35-0.75) in patients with lupus nephritis. The standardized mortality ratios of patients with and without lupus nephritis were 5.17 (95% CI 3.49-7.38) and 2.32 (95% CI 1.47-3.48), respectively. Conclusion In patients with SLE, the presence of lupus nephritis is associated with increased corticosteroid-associated damage but less corticosteroid non-associated damage. Also, mortality is significantly higher in patients with lupus nephritis than in those without lupus nephritis.

Genetic associations of LYN with systemic lupus erythematosus.

We targeted LYN, a src-tyosine kinase involved in B-cell activation, in case-control association studies using populations of European-American, African-American and Korean subjects. Our combined European-derived population, consisting of 2463 independent cases and 3131 unrelated controls, shows significant association with rs6983130 in a female-only analysis with 2254 cases and 2228 controls (P=1.1 x 10(-4), odds ratio (OR)=0.81 (95% confidence interval: 0.73-0.90)). This single nucleotide polymorphism (SNP) is located in the 5' untranslated region within the first intron near the transcription initiation site of LYN. In addition, SNPs upstream of the first exon also show weak and sporadic association in subsets of the total European-American population. Multivariate logistic regression analysis implicates rs6983130 as a protective factor for systemic lupus erythematosus (SLE) susceptibility when anti-dsDNA, anti-chromatin, anti-52 kDa Ro or anti-Sm autoantibody status were used as covariates. Subset analysis of the European-American female cases by American College of Rheumatology classification criteria shows a reduction in the risk of hematological disorder with rs6983130 compared with cases without hematological disorders (P=1.5 x 10(-3), OR=0.75 (95% CI: 0.62-0.89)). None of the 90 SNPs tested show significant association with SLE in the African American or Korean populations. These results support an association of LYN with European-derived individuals with SLE, especially within autoantibody or clinical subsets.

Genetic studies of systemic lupus erythematosus in Asia: where are we now?

There have been many genetic studies of systemic lupus erythematosus (SLE) in Asia, but the status of SLE in Asia remains unclear. Genes that have been associated with SLE in Caucasians have shown both consistent and inconsistent results in Asians. This prompted us to review studies of SLE-associated genes and compare the degree of consistency according to ethnicity in Asia. We searched PubMed and the national databases in Korea and Japan for SLE genetic studies. A total of 755 articles were found and after applying various exclusion criteria, 442 studies including 17 linkage studies, 2 genome-wide association studies and 423 candidate-gene analyses were reviewed. Nine linkage loci were confirmed to be associated with SLE susceptibility in non-Asians, but the risk locus (16q12) has been identified in only one Asian study. A total of 156 candidate genes were analyzed, of which 92 were studied in Asians. Although there were allelic (HLA-DRB1 and IRF5) or genetic heterogeneity (FCGR gene family), HLA-DRB1, the FCGR gene family, IRF5, STAT4 and MECP2 showed consistent associations with SLE susceptibility across ethnicities. In conclusion, genetic associations often vary with ethnicity, requiring validation in different ethnic groups, and hence future SLE genetic studies will require strong worldwide collaborations.

A regulatory SNP at position -899 in CDKN1A is associated with systemic lupus erythematosus and lupus nephritis.

The CDKN1A gene encoding a cell cycle inhibitor, p21(WAF1/CIP1), is located in the systemic lupus erythematosus (SLE) susceptibility locus on chromosome 6p21.2. Decreased cellular levels of p21 are associated with SLE. Here, we examine four single-nucleotide polymorphisms (SNPs) within the promoter and two in the first intron of CDKN1A for association with SLE susceptibility. A comparison of 742 Korean SLE patients with 1017 controls disclosed that one SNP (rs762624 C>A at position -899), located at a putative Myb-binding site in the promoter, was associated with SLE susceptibility (P=0.00047). This association was independent of the SLE-association signal of HLA-DRB1 on 6p21.3, as it was significant after adjustment for SLE-risk DRB1 alleles (P=0.0012). The same SNP was associated with lupus nephritis (P=0.000014). The risk allele-carrying promoter sequence displayed approximately 15% lower activity than the non-risk sequence upon fusion to the luciferase gene (P=0.025). Endogenous CDKN1A mRNA levels measured in Epstein-Barr virus-transformed B cells established from 16 control subjects were linearly correlated with a decreasing copy number of the risk allele (P=0.024). Accordingly, we conclude that the minor allele A at -899 of CDKN1A is associated with increased susceptibility to SLE and lupus nephritis, and decreased cellular levels of p21.

Specific hippocampal choline decrease in an animal model of depression.

A decreased level of the hippocampal choline signal was found in patients with depression in previous proton magnetic resonance spectroscopy ((1)H-MRS) studies. The objective of this study is to compare choline levels before and after the forced swimming test (FST), an animal model of depression typically used for assessing antidepressant activity. (1)H-MRS spectra were obtained from both the left and right hippocampus. After the FST, rats showed a significant decrease of the choline/creatine (Cho/Cr, p = 0.037) and choline/N-acetylaspartate (Cho/NAA, p = 0.048) ratios in the left hippocampus, but not in the right hippocampus. This finding was analogous to results from patients with depression. It suggests that decreased Cho/Cr and Cho/NAA ratios in the left hippocampal regions might be considered to be biomarkers in rats with depression.

Cellular regulation of bovine intramuscular adipose tissue development and composition.

It is well documented that grain feeding stimulates adipogenesis in beef cattle, whereas pasture feeding depresses the development of adipose tissues, including intramuscular (i.m.) adipose tissue. Additionally, production practices that depress adipocyte differentiation also limit the synthesis of MUFA. Marbling scores and MUFA increase in parallel suggesting that stearoyl-coenzyme A desaturase (SCD) gene expression is closely associated with and necessary for marbling adipocyte differentiation. Similarly, marbling scores and fatty acid indices of SCD activity are depressed in response to dietary vitamin A restriction. In bovine preadipocytes, vitamins A and D both decrease glycerol-3-phosphate dehydrogenase (GPDH) activity, an index of adipocyte differentiation, whereas incubation of bovine preadipocytes with l-ascorbic acid-2-phosphate increases GPDH activity. Exposing bovine preadipocytes to zinc also stimulates adipogenesis, putatively by inhibiting nitric oxide (NO) production. However, incubation of bovine preadipocytes with arginine, a biological precursor of NO, strongly promotes differentiation in concert with increased SCD expression. This suggests that the effect of either arginine or zinc on adipogenesis is independent of NO synthesis in bovine preadipocytes. Enhanced expression of SCD is associated with a greater accumulation of MUFA both in bovine preadipocyte cultures and during development in growing steers. In bovine preadipocytes, trans-10, cis-12 CLA strongly depresses adipocyte differentiation and SCD gene expression, thereby reducing MUFA concentrations. The bovine preadipocyte culture studies suggest that any production practice that elevates vitamins A or D or trans-10, cis-12 CLA in bovine adipose tissue will reduce i.m. adipose tissue development. Conversely, supplementation with vitamin C or zinc may promote the development of i.m. adipose tissue.

Comparison of plasma metabolite concentrations and enzyme activities in beef cattle raised by different feeding systems in Korea, Japan and New Zealand.

Concentrations of metabolites and immunoreactive insulin (IRI) and activities of enzymes related to energy metabolism were measured in plasma of Korean and Japanese beef cattle, which were raised by the indoor feeding system programmed to feed larger amount of roughage in their growing periods and larger amount of concentrate diet in their finishing periods (Japanese feeding system), and grazing New Zealand beef cattle. By the Japanese beef grading system, Korean and Japanese beef cattle showed high beef quality score, average grade 3.3 and 3.6, respectively. The plasma free fatty acid and lactate concentrations and lactate dehydrogenase (LDH), malate dehydrogenase (MDH) and aspartate aminotransferase (AST) activities in Korean beef cattle were significantly higher than those in Japanese beef cattle. The plasma lactate concentration in Korean beef cattle was 8.40 mmol/l, which was similar to the values observed in lactic acidosis. The higher activities of plasma LDH, MDH and AST may indicate slight liver damage by slightly acidotic conditions in Korean beef cattle. New Zealand beef cattle fed on pasture which they harvest by grazing showed significantly lower plasma glucose, cholesterol, lactate and IRI concentrations and enzyme activities than those in Korean and Japanese beef cattle fed on larger amount of concentrate diets. Plasma metabolite concentrations and energy metabolism-related enzyme activities may be good indicators for evaluating metabolic conditions of beef cattle raised by different feeding systems.

MERTK polymorphisms associated with risk of haematological disorders among Korean SLE patients.

OBJECTIVE: The MER receptor tyrosine kinase (MERTK) gene is critical for the efficient clearance of apoptotic cells and has implications for inflammation and autoimmune diseases such as systemic lupus erythematosus (SLE). We investigated the genetic polymorphisms in MERTK to evaluate it as a potential candidate gene for a host genetic study of SLE and clinical manifestations in patients with SLE. METHODS: By resequencing the coding and flanking regions of the MERTK gene in 24 unrelated Koreans, 37 polymorphisms were identified. Based on gene position, minor allele frequency and inter-single-nucleotide polymorphism (SNP) linkage disequilibrium, six of these polymorphisms were selected for subsequent genotyping and association analysis with the risk of SLE and haematological disorders in 350 Korean SLE patients and 330 controls. RESULTS: Although no significant associations with the risk of SLE were found, logistic regression analyses revealed that variants +465C > G (P = 0.05) and +130215insdelT (P = 0.0005) were significantly associated with decreased risk of leucopenia in SLE patients. Further, +465C > G, +95616G > A, +123157A > G and the haplotype ht1 also showed significant associations (P = 0.006-0.05) with a decreased risk of lymphopenia in SLE patients. CONCLUSION: Our findings suggest that polymorphisms in MERTK might be one of the genetic risk factors for presenting leucopenia and lymphopenia in SLE patients.

Lipid characteristics of subcutaneous adipose tissue and M. longissimus thoracis of Angus and Wagyu steers fed to US and Japanese endpoints.

We hypothesized that the concentrations of monounsaturated fatty acids (MUFA) and cholesterol of adipose tissue and M. longissimus thoracis would not differ between Angus and American Wagyu steers when fed to a typical US live weight, but would diverge when fed to a Japanese live weight. To test this, 8 steers of each breed type were assigned to a high-energy, corn-based diet, and another 8 steers of each breed type were fed coastal bermuda grass hay diet, supplemented with the corn-based diet to achieve a daily gain of 0.9kg/d. Targeted final body weights were 525kg for steers fed for 8 or 12mo the corn- or hay-based diets, respectively, and were 650kg for steers fed for 16 or 20mo the corn- or hay-based diets. Digesta concentrations of stearic (18:0) and trans-vaccenic acid decreased, whereas linoleic acid (18:2n-6) increased between the US and Japanese endpoints (all P⩽0.03). α-Linolenic acid (18:3n-3) increased in digesta only in the hay-fed steers during this time. Plasma concentrations of palmitic (16:0) and palmitoleic acid (16:1n-7), and the 16:1:18:0 ratio, were higher in Angus steers than in Wagyu steers. Also, the plasma 16:1:18:0 ratio was decreased by hay feeding in Angus steers, but increased in Wagyu steers, when fed to the Japanese endpoint. Concentrations of oleic (18:1n-9), linoleic, α-linolenic, and 18:2trans-10,cis-12 conjugated linoleic acid all were higher in Wagyu than in Angus subcutaneous (s.c.) adipose tissue, whereas myristic (14:0) and palmitic acid were higher in Angus s.c. adipose tissue (P⩽0.05). All MUFA increased, and saturated fatty acids decreased, between the US and Japanese endpoints. Slip points of lipids in s.c. adipose tissue were over 10°C lower (P=0.01) in Japanese-endpoint steers than in US endpoint steers, consistent with the overall increase in MUFA with time on feed. The concentration of cholesterol in the M. longissimus thoracis increased with time, which may have been related to the increase in oleic acid. Because the breed×endpoint interaction was not significant for cholesterol or any of the adipose tissue fatty acids, we conclude that our original hypothesis was incorrect. Of the three factors tested (breed type, diet, and slaughter age endpoint), endpoint had the greatest effect on adipose tissue lipid composition.

Genetic diversity of north-east Asian cattle based on microsatellite data.

In order to assess the genetic variability and population structure of north-east Asian cattle, 13 microsatellite loci were analysed for a total of 200 individuals including Korean, Chinese, Japanese Black and European Holstein cattle. Observed and expected heterozygosity, two estimators (F(ST) and G(ST)) of gene differentiation, and Nei's DA distance were evaluated. Based on expected mean heterozygosity, the lowest genetic diversity was exhibited in Japanese Black cattle (H(E)=0.471), and the highest in Chinese cattle (H(E)=0.744). Korean cattle revealed a relatively high degree of genetic diversity (H(E)=0.728). Average proportion of genetic variation because of interpopulation subdivision among north-east Asian cattle varied between 10.9 and 9.9%, depending on the estimator used. N-J tree based on Nei's DA genetic distance showed that Korean and Chinese cattle are closely related, whereas Japanese Black cattle are clearly distinct from the other two populations, forming a north-east Asian outgroup.

Modulation of expression of fos and Ha-ras oncogenes and ornithine decarboxylase activity in mammary gland and liver of young female rats by the absence of dietary lipotropes.

There is evidence that diets deficient in lipotropes [methionine, choline, pteroylmonoglutamic acid (folic acid), and cyanocobalamin (vitamin B12)] induce and enhance hepatocarcinogenesis. This research examined the extent to which dietary lipotropes modify cellular oncogene expression and ornithine decarboxylase activity in mammary gland and liver of rats. Eighteen female Sprague-Dawley rats (8 wk old) were fed 3 wk on one of three diets: 1) a control synthetic diet; 2) a methyl-deficient diet lacking choline, methionine, pteroylmonoglutamic acid, and cyanocobalamin; or 3) a diet supplemented with twice the amount of each lipotrope as in the control synthetic diet. The group fed the methyl-deficient diet gained less body weight than groups fed the control or methyl-supplemented diet. The group fed the methyl-deficient diet had approximately 5- and 11-fold greater fos transcription in mammary gland and liver, respectively, than did the control group. The expression of the Ha-ras gene in mammary gland and liver of the group fed the methyl-deficient diet was increased by 4- and 6-fold compared with that of the control. Ornithine decarboxylase activity, considered to be a developmental marker, was higher in liver and mammary gland of the group fed the methyl-deficient diet than in either the group fed control synthetic diet or the group fed the methyl-supplemented diet. The methyl-deficient diet may have caused activation of the transcription factor fos and thus the activation of the transcription regulatory complex, AP-1. In turn, AP-1 may regulate genes, such as ornithine decarboxylase, which are responsible for cell proliferation and differentiation.

Lipotrope-modified diets enhance nitrosomethylurea-induced mammary carcinogenesis in female rats.

This study was conducted to determine the effects of lipotrope-modified (deficient or supplemented) diets on nitrosomethylurea- (NMU) induced mammary tumorigenesis. Eighty female Sprague-Dawley rats (4 wks old) were assigned to one of the following groups: control-synthetic diet (CSD), containing all required lipotropes; choline-methionine-deficient diet (CMD); methyl-deficient diet (MDD), lacking all lipotropes; and methyl-supplemented diet (MSD), containing twice as much of each lipotrope as the CSD diet. All animals were injected with NMU after a three-week dietary treatment period. MDD and MSD groups had shorter tumor latency periods (73 and 74 days, respectively) than the CSD group (105 days). Number of tumors per rat was significantly increased in the MDD group (4.6) compared with CSD (1.6), CMD (2.1), and MSD (2.5) groups. The results indicate that dietary manipulation of lipotropes in young female rats enhanced NMU-induced mammary tumorigenesis.

Developmental stages and energy restriction affect cellular oncogene expression in tissues of female rats.

The influence of developmental stages and energy restriction on c-Ha-ras and c-fos cellular oncogene mRNA level was examined in mammary tissue and liver of female rats. c-Ha-ras and c-fos RNA transcripts in mammary tissue peaked at 9 wk of age, decreased during pregnancy, were lowest at mid-lactation and then increased at 25 wk. c-Ha-ras in liver showed highest expression at 5 wk and was hardly detectable at 11 wk, mid-pregnancy, mid-lactation and 25 wk. Liver c-fos and c-Ha-ras mRNA levels were similar. Lower c-Ha-ras and c-fos mRNA levels were observed in mammary tissue of energy-restricted (to 70% of ad libitum intake) rats at 25 wk compared with those allowed ad libitum access to food. Energy restriction did not affect liver c-Ha-ras and c-fos mRNA levels throughout the experimental period. These results demonstrate that developmental stages and energy restriction affect cellular oncogene mRNA levels with tissue-specific patterns.

Energy restriction and testosterone implants alter ornithine decarboxylase gene expression in kidney of female rats.

Effects of testosterone and energy restriction (30%) on enzyme activity and mRNA level of ornithine decarboxylase (ODC) were studied in 19-wk-old female Sprague-Dawley rats. Testosterone implantation for 1 wk elicited a sixfold increase in ODC activity and a fourfold increase in ODC mRNA transcripts in the kidney. Energy restriction also increased renal ODC activity. Further, the 56% increase in ODC enzyme activity observed in the testosterone-implanted, energy-restricted group was greater than the 24% increase in the placebo-implanted, energy-restricted group. No changes in renal ODC mRNA levels were observed in the energy-restricted groups. These observations suggest that translational or post-translational mechanism(s) are involved in the greater renal ODC activity in energy-restricted rats.