Hiroaki Hamada

Therapeutic efficacy of intra-articular adrenomedullin injection in antigen-induced arthritis in rabbits

IntroductionAdrenomedullin is a potent vasodilatory and hypotensive peptide as well as an endogenous immunomodulatory factor with predominantly anti-inflammatory effects. The purpose of the present study was to evaluate the therapeutic effects of adrenomedullin in rabbits with antigen-induced arthritis, an experimental model of rheumatoid arthritis.MethodsFollowing the induction of arthritis in both knee joints by ovalbumin injection into the joint spaces of pre-immunized rabbits, increasing daily doses of adrenomedullin were injected into the knee joint spaces or saline was injected into the contralateral knee joint spaces as the control. For time-course experiments, adrenomedullin and saline were injected into the knee joint spaces daily for 7 days and 20 days. The degree of joint swelling and the histological change in the knee joints injected with adrenomedullin were compared with the control knee joints. Histological evaluation of the infrapatellar fat pads and synovial tissue was performed. TNFα, IL-6, vascular endothelial growth factor and transforming growth factor-beta mRNA levels in the synovial tissue were measured using real-time quantitative PCR.ResultsDaily injections of adrenomedullin into the knee joint spaces of rabbits with antigen-induced arthritis decreased joint swelling. Histological examination revealed that adrenomedullin reduced edematous changes and the infiltration of inflammatory cells in the synovial tissues. Analysis of mRNA levels showed that adrenomedullin significantly reduced TNFα mRNA expression by 21% to 49% in a dose-dependent manner, and dose-dependently increased IL-6 mRNA expression by 45% to 121%.ConclusionsThese results suggest that daily injections of adrenomedullin into the knee joint spaces of rabbits with antigen-induced arthritis ameliorated the inflammatory response in arthritic joints. Adrenomedullin may thus be useful as a treatment for rheumatoid arthritis; however, the effect of adrenomedullin on IL-6 production in the synovial tissue may be an undesirable adverse effect in rheumatoid arthritis therapy.

Expression of adrenomedullin and its receptor by chondrocyte phenotype cells.

For clarifying a process of de-differentiation in culturing chondrocytes, the present study was undertaken to investigate the secretion of adrenomedullin (AM) by chondrocyte phenotype cells and whether or not AM effects this proliferation in a cAMP-dependent fashion. Chondrocyte phenotype cells expressed AM and the AM receptor, and secreted high concentration of AM into the culture medium. When added to cultures, AM increased the intracellular cAMP level and decreased the number of these cells in a similar concentration-dependent fashion. Addition of forskolin and dibutyryl-cAMP caused a significant decrease in the number of these cells. Furthermore, the effect of AM was inhibited by a cAMP-dependent protein kinase A inhibitor (H89). The present findings indicate that AM has an autocrine/paracrine type of anti-proliferative effect on these cells mediated via a cAMP-dependent pathway and raise the possibility that AM plays a role in the local modulation of a process of de-differentiation by culturing chondrocyte phenotype cells.

Segmental copy number loss in the region of Semaphorin 4D gene in patients with acetabular dysplasia.

Acetabular dysplasia (AD) appears to be a multi‐factorial disease, which may involve both genetic and environmental factors and whose pathogenesis remains obscure. The present study aims to identify a genetic variation that might confer risk of AD. We performed whole‐genome screening of a copy number variation (CNV) using a deCODE‐Illumina CNV beadchip with 20 female AD patients and 131 control subjects. Subsequently, Agilents region‐targeted high‐density oligonucleotide tiling microarray was used to analyze 64 female AD patients and 32 female control subjects. By sequential analyses, we found a copy number loss in 18 of 64 AD patients, but none in the 32 controls. The loss occurred within a 472 kb region on 9q22.2, which harbors the gene for Semaphorin 4D (Sema4D; 18/64 vs. 0/32, p = 4.81 × 10−4, OR = 25.86). We suggest that a copy number loss of the Sema4D gene region may play a role in the etiology of AD.

Copy number loss in the region of the ASPN gene in patients with acetabular dysplasia

Objectives We have previously investigated an association between the genome copy number variation (CNV) and acetabular dysplasia (AD). Hip osteoarthritis is associated with a genetic polymorphism in the aspartic acid repeat in the N-terminal region of the asporin (ASPN) gene; therefore, the present study aimed to investigate whether the CNV of ASPN is involved in the pathogenesis of AD. Methods Acetabular coverage of all subjects was evaluated using radiological findings (Sharp angle, centre-edge (CE) angle, acetabular roof obliquity (ARO) angle, and minimum joint space width). Genomic DNA was extracted from peripheral blood leukocytes. Agilent’s region-targeted high-density oligonucleotide tiling microarray was used to analyse 64 female AD patients and 32 female control subjects. All statistical analyses were performed using EZR software (Fisher’s exact probability test, Pearson’s correlation test, and Student’s t-test). Results CNV analysis of the ASPN gene revealed a copy number loss in significantly more AD patients (9/64) than control subjects (0/32; p = 0.0212). This loss occurred within a 60 kb region on 9q22.31, which harbours the gene for ASPN. The mean radiological parameters of these AD patients were significantly worse than those of the other subjects (Sharp angle, p = 0.0056; CE angle, p = 0.0076; ARO angle, p = 0.0065), and all nine patients required operative therapy such as total hip arthroplasty or pelvic osteotomy. Moreover, six of these nine patients had a history of operative or conservative therapy for developmental dysplasia of the hip. Conclusions Copy number loss within the region harbouring the ASPN gene on 9q22.31 is associated with severe AD. A copy number loss in the ASPN gene region may play a role in the aetiology of severe AD. Cite this article: T. Sekimoto, M. Ishii, M. Emi, S. Kurogi, T. Funamoto, Y. Yonezawa, T. Tajima, T. Sakamoto, H. Hamada, E. Chosa. Copy number loss in the region of the ASPN gene in patients with acetabular dysplasia: ASPN CNV in acetabular dysplasia. Bone Joint Res 2017;6:439–445. DOI: 10.1302/2046-3758.67.BJR-2016-0094.R1.

Possible association of single nucleotide polymorphisms in the 3' untranslated region of HOXB9 with acetabular overcoverage

Objectives Excessive acetabular coverage is the most common cause of pincer-type femoroacetabular impingement. To date, an association between acetabular over-coverage and genetic variations has not been studied. In this study we investigated the association between single nucleotide polymorphisms (SNPs) of paralogous Homeobox (HOX)9 genes and acetabular coverage in Japanese individuals to identify a possible genetic variation associated with acetabular over-coverage. Methods We investigated 19 total SNPs in the four HOX9 paralogs, then focused in detail on seven of those located in the 3’ untranslated region of HOXB9 (rs8844, rs3826541, rs3826540, rs7405887, rs2303485, rs2303486, rs79931349) using a case-control association study. The seven HOXB9 SNPs were genotyped in 316 subjects who had all undergone radiological examination. The association study was performed by both single-locus and haplotype-based analyses. Results The genotype and allele frequencies of the five HOXB9 SNPs showed significant association with acetabular over-coverage compared with controls (rs7405887 OR = 3.16, p = 5.29E-6, 95% CI 1.91 to 5.25). A significant difference was also detected when haplotypes were evaluated (OR = 2.59, p = 2.61E-5, 95% CI 1.65 to 4.08). The two HOXB9 SNPs (rs2303485, rs2303486) were associated with decreased acetabular coverage (rs2303485 OR = 0.524, p = 0.0091, 95% CI 0.322 to 0.855; rs2303486 OR = 0.519, p = 0.011, 95% CI 0.312 to 0.865). Conclusions The five HOXB9 SNPs (rs8844, rs3826541, rs3826540, rs7405887, rs79931349) were associated with acetabular over-coverage. On the other hand, the two SNPs (rs2303485 and rs2303486) were associated with the lower acetabular coverage. The association of rs2303486 would be consistent with the previous study. Therefore, the HOXB9 SNPs might be involved in the morphogenesis of acetabular coverage, and could be an independent risk factor for developing pincer-type femoroacetabular impingement. Cite this article: Bone Joint Res 2015;4:50–5.

Prevalence of chronic kidney disease and administration of RA-related drugs in patients with RA: The NinJa 2012 study in Japan

Abstract Objectives: To estimate the prevalence of chronic kidney disease in patients with rheumatoid arthritis (RA) and the administration of disease-modifying anti-rheumatic-drugs (DMARDs), using data from the National Database of Rheumatic Disease by iR-net in Japan (NinJa) 2012 study. Methods: From a total of 11,940 RA patients, 7135 who underwent an estimated glomerular filtration rate (eGFR) test were studied. Renal dysfunction staging was assessed using Japanese eGFR equations and classified according to the Kidney Disease Improving Global Outcomes 2012 clinical practice guideline. Results: The prevalence of GFR stages was as follows: stage G1, 25.4%; stage G2, 55.9%; stage G3, 17.5%; stage G4, 0.8%; and stage G5, 0.2%. Overall, 92.7% of patients received at least one DMARD. Sulfasalazine, tacrolimus, and biologics (except inflixmab) were administered in all GFR stages. Methotrexate was not prescribed in patients with stage G5, but methotrexate 3.5 mg/week (mean) was prescribed in four patients (6.8%) with stage G4. Non-steroidal anti-inflammatory drugs and glucocorticoids were prescribed in 40.5% and 43.7% of patients, respectively. Conclusion: The prevalence of kidney disease in this large sample of RA patients was higher than that in the general population, and the results suggest that RA patients with renal dysfunction require careful drug selection.

Pathological Renal Findings of Chronic Renal Failure in a Patient with the E66Q Mutation in the α-galactosidase A Gene

A 66-year-old Japanese man was diagnosed with interstitial nephritis on a renal biopsy at 45 years of age and began to receive hemodialysis at 65 years of age. He was suspected of having Fabry disease as a result of a screening study for Fabry disease performed in hemodialysis patients. He had an E66Q mutation in the α-galactosidase A gene. We conducted an electron microscopic examination of a renal biopsy specimen obtained when the patient was diagnosed with chronic renal failure at 45 years of age in order to elucidate the pathogenicity of the E66Q mutation. Interestingly, an electron microscopic examination of the renal biopsy specimen indicated no characteristic findings of Fabry disease.

Finite element analysis of the tibial bone graft in cementless total knee arthroplasty

BackgroundAchieving stability of the tibial implant is essential following cementless total knee arthroplasty with bone grafting. We investigated the effects of bone grafting on the relative micromotion of the tibial implant and stress between the tibial implant and adjacent bone in the immediate postoperative period.MethodsTibial implant models were developed using a nonlinear, three-dimensional, finite element method. On the basis of a preprepared template, several bone graft models of varying sizes and material properties were prepared.ResultsMicromotion was larger in the bone graft models than in the intact model. Maximum micromotion and excessive stress in the area adjacent to the bone graft were observed for the soft and large graft models. With hard bone grafting, increased load transfer and decreased micromotion were observed.ConclusionsAvoidance of large soft bone grafts and use of hard bone grafting effectively reduced micromotion and undue stress in the adjacent area.

Aortic aneurysm as a complication of myeloperoxidase-antineutrophil cytoplasmic antibody-associated vasculitis

Abstract Myeloperoxidase-antineutrophil cytoplasmic antibody-associated vasculitis (MPO-AAV) does not usually involve large vessels, such as the aorta. However, we experienced three cases having an aortic aneurysm as a complication of MPO-AAV with renal insufficiency. In one patient it involved the onset of descending aortic dissection during treatment for MPO-AAV; another two patients had an abdominal aortic aneurysm at the time of our diagnosis of MPO-AAV. Although we found no pathological evidence in our patients, MPO-AAV might result in large vessel inflammation. Therefore, we suggest that patients with MPO-AAV should be examined by computed tomography scan to check for the presence of an aortic aneurysm.

Role of Adrenomedullin in Patients with Rheumatoid Arthritis

The mechanism of arthritis including rheumatoid arthritis (RA) has not been clarified enough in terms of its molecular biology. However, owing to advances in techniques to produce monoclonal antibodies and genetic engineering techniques to produce recombinant proteins, targeted therapies have progressed. Tumor necrosis factor-alpha (TNF-), interleukin-1 (IL-1), interleukin-6 (IL-6), and T-cells (CD28) are representative targets, and therapies targeting them have produced epoch-making results in the treatment of RA. In this text, we report the role and effects of therapy with adrenomedullin as a novel, promising targeted therapy. Adrenomedullin (ADM) is a peptide that was discovered by the assay-based monitoring of platelet cAMP activity (Kitamura et al., 1993). This protein shows not only marked vasodilatory but also in vivo and in vitro anti-inflammatory activities (Isumi et al., 1998). To determine the efficacy of ADM in patients with RA, we researched the plasma level of ADM and its correlation with the plasma CRP level. The result showed that the plasma ADM level in patients with RA was twice that in healthy controls and strongly correlated with the blood CRP level, and the ADM level in synovial tissue in patients with RA was 3 times that in patients with osteoarthritis (Chosa et al., 2003). Then, thorough research is needed to examine whether the cultured RA synoviocytes are related to the ADM in vitro. The result showed that ADM was produced from RA synoviocytes and reduced the IL-6 level in cultured media (Fig. 5). These results suggest the close involvement of ADM in arthritis in RA. Therefore, to examine the therapeutic efficacy of intra-articular ADM injection, we injected ADM into the joint of a rabbit model of adjuvant arthritis, and confirmed the inhibition of arthritis at an early stage. These findings suggest the potentiality of ADM as a novel drug for the treatment of arthritis in RA (Okura et al., 2008).