JMJD6 Autoantibodies as a Potential Biomarker for Inflammation-Related Diseases.

Inflammation is closely associated with cerebrovascular diseases, cardiovascular diseases, diabetes, and cancers, and it is accompanied by the development of autoantibodies in the early stage of inflammation-related diseases. Hence, it is meaningful to discover novel antibody biomarkers targeting inflammation-related diseases. In this study, Jumonji C-domain-containing 6 (JMJD6) was identified by the serological identification of antigens through recombinant cDNA expression cloning. In particular, JMJD6 is an antigen recognized in serum IgG from patients with unstable angina pectoris (a cardiovascular disease). Then, the serum antibody levels were examined using an amplified luminescent proximity homogeneous assay-linked immunosorbent assay and a purified recombinant JMJD6 protein as an antigen. We observed elevated levels of serum anti-JMJD6 antibodies (s-JMJD6-Abs) in patients with inflammation-related diseases such as ischemic stroke, acute myocardial infarction (AMI), diabetes mellitus (DM), and cancers (including esophageal cancer, EC; gastric cancer; lung cancer; and mammary cancer), compared with the levels in healthy donors. The s-JMJD6-Ab levels were closely associated with some inflammation indicators, such as C-reactive protein and intima-media thickness (an atherosclerosis index). A better postoperative survival status of patients with EC was observed in the JMJD6-Ab-positive group than in the negative group. An immunohistochemical analysis showed that JMJD6 was highly expressed in the inflamed mucosa of esophageal tissues, esophageal carcinoma tissues, and atherosclerotic plaques. Hence, JMJD6 autoantibodies may reflect inflammation, thereby serving as a potential biomarker for diagnosing specific inflammation-related diseases, including stroke, AMI, DM, and cancers, and for prediction of the prognosis in patients with EC.

Serum anti-PCK1 antibody levels are a prognostic factor for patients with diabetes mellitus.

BACKGROUND: Autoantibodies develop in autoimmune diseases, cancer, diabetes mellitus (DM), and atherosclerosis-related diseases. However, autoantibody biomarkers have not been successfully examined for diagnosis and therapy. METHODS: Serological identification of antigens through recombinant cDNA expression cloning (SEREX) was used for primary screening of antigens. The cDNA product was expressed in bacteria and purified. Amplified luminescent proximity homogeneous assay-linked immunosorbent assay (AlphaLISA) was used to evaluate antibody levels in serum samples. RESULTS: Phosphoenolpyruvate carboxykinase 1 (PCK1) was recognized as an antigen by serum IgG antibodies in the sera of patients with atherosclerosis. AlphaLISA showed significantly higher serum antibody levels against recombinant PCK1 protein in patients with DM and cardiovascular disease than in healthy donors, but not in those with acute ischemic stroke, transient ischemic attack, or obstructive sleep apnea syndrome. The area under the receiver operating characteristic curve for anti-PCK1 antibodies was 0.7024 for DM. The serum anti-PCK1 antibody levels were associated with age, platelet count, and blood pressure. Anti-PCK1-antibody-positive patients showed significantly lower overall survival than the negative patients. CONCLUSIONS: Serum anti-PCK1 antibody levels were found to be associated with DM. The anti-PCK1 antibody marker is useful for predicting the overall survival of patients with DM.

Targeted long-read sequencing identifies missing pathogenic variants in unsolved Werner syndrome cases.

BACKGROUND: Werner syndrome (WS) is an autosomal recessive progeroid syndrome caused by variants in WRN. The International Registry of Werner Syndrome has identified biallelic pathogenic variants in 179/188 cases of classical WS. In the remaining nine cases, only one heterozygous pathogenic variant has been identified. METHODS: Targeted long-read sequencing (T-LRS) on an Oxford Nanopore platform was used to search for a second pathogenic variant in WRN. Previously, T-LRS was successfully used to identify missing variants and analyse complex rearrangements. RESULTS: We identified a second pathogenic variant in eight of nine unsolved WS cases. In five cases, T-LRS identified intronic splice variants that were confirmed by either RT-PCR or exon trapping to affect splicing; in one case, T-LRS identified a 339 kbp deletion, and in two cases, pathogenic missense variants. Phasing of long reads predicted all newly identified variants were on a different haplotype than the previously known variant. Finally, in one case, RT-PCR previously identified skipping of exon 20; however, T-LRS did not detect a pathogenic DNA sequence variant. CONCLUSION: T-LRS is an effective method for identifying missing pathogenic variants. Although limitations with computational prediction algorithms can hinder the interpretation of variants, T-LRS is particularly effective in identifying intronic variants.

Iron overload may be critical for liver dysfunction in anorexia nervosa, and the role of haematocrit-adjusted albumin in assessing nutritional status: a case report.

BACKGROUND: Anorexia nervosa (AN) is frequently associated with liver dysfunction, but the precise mechanism remains undefined. Since the nutritional marker albumin has a low correlation with changes in body weight in AN, and patients with AN often have dehydration as a complication, we also examined whether haematocrit (HCT)-adjusted serum albumin could be a better nutritional marker in AN. CASE PRESENTATION: We describe a 15-year-old girl with severe weight loss and liver damage whose liver enzymes normalized after 1.5 months of hospitalization and weight gain. We found a significant correlation between body weight (BW) and HCT-adjusted serum albumin (Spearman's rank correlation coefficient (rs) = 0.66, P = 5.28 × 10-3) and between BW and alanine aminotransferase (ALT) (rs = -0.825, P = 8.45 × 10-5). After division by HCT, correlations between serum albumin and ALT (rs = -0.835, P = 5.24 × 10-5) and between the iron-storage protein ferritin and the liver enzyme gamma-glutamyl transferase (rs = 1.0, P = 0.017) were also statistically significant. CONCLUSION: These results suggest that improvement of the nutritional status in AN could relieve liver dysfunction and facilitate iron transport. Since a decrease in the iron-transport protein transferrin presumably increases labile non-transferrin-bound iron, resulting in excess reactive oxygen species production, a defect in iron transport due to malnutrition could be one of the causes of liver injury in AN. In addition, HCT-adjusted albumin could be a better marker than its raw data to assess changes in nutritional status in AN.

Rothmund-Thomson syndrome investigated by two nationwide surveys in Japan.

BACKGROUND: Rothmund-Thomson syndrome (RTS) is an autosomal recessive genetic disorder characterized by poikiloderma of the face, small stature, sparse scalp hair, juvenile cataract, radial aplasia, and predisposition to cancers. Due to the rarity of RTS, the situation of patients with RTS in Japan has not been elucidated. METHODS: In 2010 and 2020, following the results of a primary questionnaire survey, a secondary questionnaire survey on RTS was conducted nationwide to investigate the number of RTS cases and their associated skin lesions, bone lesions, other clinical features, and quality of life in Japan. RESULTS: In 2010 and 2020, 10 and eight patients with RTS were recruited, respectively. Skin lesions such as poikiloderma, erythema, pigmentation, and abnormal scalp hair were observed in almost all cases. Bone lesions were observed in four cases in the 2010 and 2020 surveys, respectively. Two cases had mutations in the RECQL4 gene in the 2020 survey. CONCLUSIONS: Two nationwide surveys have shown the actual situation of patients with RTS in Japan. Cutaneous and bone manifestations are important for the diagnosis of RTS. However, many patients have no RECQL4 mutations. The novel causative gene of RTS should be further elucidated.

Serum anti-DIDO1, anti-CPSF2, and anti-FOXJ2 antibodies as predictive risk markers for acute ischemic stroke.

BACKGROUND: Acute ischemic stroke (AIS) is a serious cause of mortality and disability. AIS is a serious cause of mortality and disability. Early diagnosis of atherosclerosis, which is the major cause of AIS, allows therapeutic intervention before the onset, leading to prevention of AIS. METHODS: Serological identification by cDNA expression cDNA libraries and the protein array method were used for the screening of antigens recognized by serum IgG antibodies in patients with atherosclerosis. Recombinant proteins or synthetic peptides derived from candidate antigens were used as antigens to compare serum IgG levels between healthy donors (HDs) and patients with atherosclerosis-related disease using the amplified luminescent proximity homogeneous assay-linked immunosorbent assay. RESULTS: The first screening using the protein array method identified death-inducer obliterator 1 (DIDO1), forkhead box J2 (FOXJ2), and cleavage and polyadenylation specificity factor (CPSF2) as the target antigens of serum IgG antibodies in patients with AIS. Then, we prepared various antigens including glutathione S-transferase-fused DIDO1 protein as well as peptides of the amino acids 297-311 of DIDO1, 426-440 of FOXJ2, and 607-621 of CPSF2 to examine serum antibody levels. Compared with HDs, a significant increase in antibody levels of the DIDO1 protein and peptide in patients with AIS, transient ischemic attack (TIA), and chronic kidney disease (CKD) but not in those with acute myocardial infarction and diabetes mellitus (DM). Serum anti-FOXJ2 antibody levels were elevated in most patients with atherosclerosis-related diseases, whereas serum anti-CPSF2 antibody levels were associated with AIS, TIA, and DM. Receiver operating characteristic curves showed that serum DIDO1 antibody levels were highly associated with CKD, and correlation analysis revealed that serum anti-FOXJ2 antibody levels were associated with hypertension. A prospective case-control study on ischemic stroke verified that the serum antibody levels of the DIDO1 protein and DIDO1, FOXJ2, and CPSF2 peptides showed significantly higher odds ratios with a risk of AIS in patients with the highest quartile than in those with the lowest quartile, indicating that these antibody markers are useful as risk factors for AIS. CONCLUSIONS: Serum antibody levels of DIDO1, FOXJ2, and CPSF2 are useful in predicting the onset of atherosclerosis-related AIS caused by kidney failure, hypertension, and DM, respectively.

R3hdml regulates satellite cell proliferation and differentiation.

In this study, we identified a previously uncharacterized skeletal satellite cell-secreted protein, R3h domain containing-like (R3hdml). Expression of R3hdml increases during skeletal muscle development and differentiation in mice. Body weight and skeletal muscle mass of R3hdml knockout (KO) mice are lower compared to control mice. Expression levels of cell cycle-related markers, phosphorylation of Akt, and expression of insulin-like growth factor within the skeletal muscle are reduced in R3hdml KO mice compared to control mice. Expression of R3hdml increases during muscle regeneration in response to cardiotoxin (CTX)-induced muscle injury. Recovery of handgrip strength after CTX injection was significantly impaired in R3hdml KO mice, which is rescued by R3hdml. Our results indicate that R3hdml is required for skeletal muscle development, regeneration, and, in particular, satellite cell proliferation and differentiation.

Low dose red yeast rice with monacolin K lowers LDL cholesterol and blood pressure in Japanese with mild dyslipidemia: A multicenter, randomized trial.

BACKGROUND AND OBJECTIVES: Red yeast rice contains monacolin K, an inhibitor of cholesterol synthesis, and gamma-aminobutyric acid, a neurotransmitter. The daily dose of red yeast rice and monacolin K in previous studies was relatively high; therefore, there were safety concerns. We aimed to examine the effects of low daily dose red yeast rice on arteriosclerosis in patients with mild dyslipidemia. METHODS AND STUDY DESIGN: Eighteen patients without known cardiovascular disease and unsatisfactory low-density lipoprotein cholesterol (3.96±0.19 mmol/L) controlled only by diet therapy were randomly allocated to receive low dose red yeast rice (200 mg/day) containing 2 mg monacolin K or diet therapy alone for 8 weeks. The primary outcome was the absolute change in low-density lipoprotein cholesterol. Secondary outcomes included total cholesterol, apolipoprotein B, and blood pressure. RESULTS: Low-density lipoprotein cholesterol decreased significantly in the red yeast rice group than in the diet therapy group (median [interquartile range]: control -0.20 [-0.62, 1.19] mmol/L vs. red yeast rice -0.96 [-1.05, -0.34] mmol/L, p=0.030). The red yeast rice group also exhibited significant decreases in total cholesterol, apolipoprotein B, and blood pressure. No severe treatment-related adverse effects on muscles, liver, or renal function were observed. CONCLUSIONS: We found that patients in the red yeast rice group exhibited significant reductions in lowdensity lipoprotein cholesterol, total cholesterol, apolipoprotein B, and blood pressure without any recognised adverse effect. This suggests that low daily dose red yeast rice could reduce cardiovascular risk in patients with dyslipidemia.

Serum anti-LRPAP1 is a common biomarker for digestive organ cancers and atherosclerotic diseases.

Some cancers are related to atherosclerotic diseases; therefore, these two types of disease may share some antibody biomarkers in common. To investigate this, a first screening of sera was performed from patients with esophageal squamous cell carcinoma (ESCC) or acute ischemic stroke (AIS) for serological identification of antigens using recombinant cDNA expression cloning (SEREX). The amplified luminescent proximity homogeneous assay-linked immunosorbent assay (AlphaLISA) method, which incorporates glutathione donor beads and anti-human IgG acceptor beads, was used to evaluate serum antibody levels. SEREX screening identified low-density lipoprotein receptor-related protein-associated protein 1 (LRPAP1) as a target antigen of serum IgG antibodies in the sera of patients with ESCC or AIS. Antigens, including recombinant glutathione S-transferase-fused LRPAP1 protein, were prepared to examine serum antibody levels. AlphaLISA revealed significantly higher antibody levels against the LRPAP1 protein in patients with solid cancers such as ESCC and colorectal carcinoma and some atherosclerosis-related diseases such as AIS and diabetes mellitus compared with healthy donors. Correlation analysis revealed that the elevated serum antibody levels against LRPAP1 were associated with smoking, a well-known risk factor for both cancer and atherosclerosis. Serum LRPAP1 antibody is therefore a common marker for the early diagnosis of some cancers and atherosclerotic diseases and may reflect diseases caused by habitual smoking.

Comparing the effects of ipragliflozin versus metformin on visceral fat reduction and metabolic dysfunction in Japanese patients with type 2 diabetes treated with sitagliptin: A prospective, multicentre, open-label, blinded-endpoint, randomized controlled study (PRIME-V study).

A prospective, multicentre, open-label, blinded-endpoint, randomized controlled study was conducted to evaluate the efficacy of treatment with ipragliflozin (sodium-dependent glucose transporter-2 inhibitor) versus metformin for visceral fat reduction and glycaemic control among Japanese patients with type 2 diabetes treated with sitagliptin, HbA1c levels of 7%-10%, and body mass index (BMI) ≥ 22 kg/m2 . Patients were randomly assigned (1:1) to receive ipragliflozin 50 mg or metformin 1000-1500 mg daily. The primary outcome was change in visceral fat area as measured by computed tomography after 24 weeks of therapy. The secondary outcomes were effects on glucose metabolism and lipid metabolism. Mean percentage reduction in visceral fat area was significantly greater in the ipragliflozin group than in the metformin group (-12.06% vs. -3.65%, P = 0.040). Ipragliflozin also significantly reduced BMI, subcutaneous fat area, waist circumference, fasting insulin, and homeostatic model assessment (HOMA)-resistance, and increased HDL-cholesterol levels. Metformin significantly reduced HbA1c and LDL-cholesterol levels and increased HOMA-beta. There were no severe adverse events. The use of ipragliflozin or metformin in combination with dipeptidyl peptidase-4 inhibitors, widely used in Japan, may have beneficial effects in ameliorating multiple cardiovascular risk factors.