Detecting and exploring kidney-derived extracellular vesicles in plasma.

BACKGROUND: Extracellular vesicles (EVs) have received considerable attention as ideal biomarkers for kidney diseases. Most reports have focused on urinary EVs, that are mainly derived from the cells in the urinary tract. However, the detection and the application of kidney-derived EVs in plasma remains uncertain. METHODS: We examined the kidney-derived small EVs (sEVs) in plasma that were supposedly released from renal mesangial and glomerular endothelial cells, using clinical samples from healthy controls and patients with kidney transplants. Plasma from healthy controls underwent ultracentrifugation, followed by on-bead flow cytometry, targeting α8 integrin, an antigen-specific to mesangial cells. To confirm the presence of kidney-derived sEVs in peripheral blood, plasma from ABO-incompatible kidney transplant recipients was ultracentrifuged, followed by western blotting for donor blood type antigens. RESULTS: Immunohistochemistry and immunoelectron microscopy confirmed α8 integrin expression in kidney mesangial cells and their sEVs. The CD9-α8 integrin double-positive sEVs were successfully detected using on-bead flow cytometry. Western blot analysis further revealed transplanted kidney-derived sEVs containing blood type B antigens in non-blood type B recipients, who had received kidneys from blood type B donors. Notably, a patient experiencing graft kidney loss exhibited diminished signals of sEVs containing donor blood type antigens. CONCLUSION: Our findings demonstrate the potential usefulness of kidney-derived sEVs in plasma in future research for kidney diseases.

Network Regulation of microRNA Biogenesis and Target Interaction.

MicroRNAs (miRNAs) are versatile, post-transcriptional regulators of gene expression. Canonical miRNAs are generated through the two-step DROSHA- and DICER-mediated processing of primary miRNA (pri-miRNA) transcripts with optimal or suboptimal features for DROSHA and DICER cleavage and loading into Argonaute (AGO) proteins, whereas multiple hairpin-structured RNAs are encoded in the genome and could be a source of non-canonical miRNAs. Recent advances in miRNA biogenesis research have revealed details of the structural basis of miRNA processing and cluster assistance mechanisms that facilitate the processing of suboptimal hairpins encoded together with optimal hairpins in polycistronic pri-miRNAs. In addition, a deeper investigation of miRNA-target interaction has provided insights into the complexity of target recognition with distinct outcomes, including target-mediated miRNA degradation (TDMD) and cooperation in target regulation by multiple miRNAs. Therefore, the coordinated or network regulation of both miRNA biogenesis and miRNA-target interaction is prevalent in miRNA biology. Alongside recent advances in the mechanistic investigation of miRNA functions, this review summarizes recent findings regarding the ordered regulation of miRNA biogenesis and miRNA-target interaction.

Systematic characterization of seed overlap microRNA cotargeting associated with lupus pathogenesis.

BACKGROUND: Combinatorial gene regulation by multiple microRNAs (miRNAs) is widespread and closely spaced target sites often act cooperatively to achieve stronger repression ("neighborhood" miRNA cotargeting). While miRNA cotarget sites are suggested to be more conserved and implicated in developmental control, the pathological significance of miRNA cotargeting remains elusive. RESULTS: Here, we report the pathogenic impacts of combinatorial miRNA regulation on inflammation in systemic lupus erythematosus (SLE). In the SLE mouse model, we identified the downregulation of two miRNAs, miR-128 and miR-148a, by TLR7 stimulation in plasmacytoid dendritic cells. Functional analyses using human cell lines demonstrated that miR-128 and miR-148a additively target KLF4 via extensively overlapping target sites ("seed overlap" miRNA cotargeting) and suppress the inflammatory responses. At the transcriptome level, "seed overlap" miRNA cotargeting increases susceptibility to downregulation by two miRNAs, consistent with additive but not cooperative recruitment of two miRNAs. Systematic characterization further revealed that extensive "seed overlap" is a prevalent feature among broadly conserved miRNAs. Highly conserved target sites of broadly conserved miRNAs are largely divided into two classes-those conserved among eutherian mammals and from human to Coelacanth, and the latter, including KLF4-cotargeting sites, has a stronger association with both "seed overlap" and "neighborhood" miRNA cotargeting. Furthermore, a deeply conserved miRNA target class has a higher probability of haplo-insufficient genes. CONCLUSIONS: Our study collectively suggests the complexity of distinct modes of miRNA cotargeting and the importance of their perturbations in human diseases.

Mesenchymal stem cells exert renoprotection via extracellular vesicle-mediated modulation of M2 macrophages and spleen-kidney network.

Adipose-derived mesenchymal stem cells (ASCs) have shown therapeutic potentials against refractory diseases. However, the detailed therapeutic mechanisms remain unclear. Here, we report the therapeutic actions of human ASCs in nephritis, focusing on cellular dynamics and multi-organ networks. Intravenously-administered ASCs accumulated in spleen but not kidneys. Nevertheless, ASCs increased M2 macrophages and Tregs in kidneys and drove strong renoprotection. Splenectomy abolished these therapeutic effects. ASC-derived extracellular vesicles (EVs) were transferred to M2 macrophages, which entered the bloodstream from spleen. EVs induced the transcriptomic signatures of hyperpolarization and PGE2 stimulation in M2 macrophages and ameliorated glomerulonephritis. ASCs, ASC-derived EVs, and EV-transferred M2 macrophages enhanced Treg induction. These findings suggest that EV transfer from spleen-accumulated ASCs to M2 macrophages and subsequent modulation of renal immune-environment underlie the renoprotective effects of ASCs. Our results provide insights into the therapeutic actions of ASCs, focusing on EV-mediated modulation of macrophages and the spleen-kidney immune network.

Association between conscientiousness and peritoneal dialysis-related peritonitis.

AIM: Peritonitis is a critical complication in patients undergoing peritoneal dialysis. Patient characteristics, especially conscientiousness, potentially influence self-care activity. This study aimed to examine the relationship between the peritonitis risk and the patient conscientiousness. METHODS: This retrospective observational study was conducted at three Japanese medical centres in 2018. Adult patients undergoing peritoneal dialysis were evaluated for conscientiousness with the Japanese version of the Ten-Item Personality Inventory. We followed the patients from peritoneal dialysis induction retrospectively on chart review. The primary outcome was time to first episode of peritonitis after initiation of peritoneal dialysis in the entire observation period. Hazard ratios were estimated using a Cox model adjusted for potential confounders. RESULTS: Among 111 patients enrolled, 32 experienced peritonitis. The patients were divided into the more conscientious group (n = 64) and less conscientious group (n = 47) based on a mean conscientiousness score of 3.991. Additionally, less conscientious patients tended to be at higher risk for peritonitis during the entire study period (hazard ratio adjusted for age and sex, 1.99; 95% confidence interval, 0.98-4.03; p = .057). The increased risk in the less conscientious group was statistically significant in the first year (adjusted hazard ratio, 6.98; 95% confidence interval, 1.42-34.40; p = .017). CONCLUSION: Less conscientious patients significantly have shorter peritonitis-free survival in the early phase after peritoneal dialysis induction although not significantly during the entire study period. Personality trait assessment using the Ten-Item Personality Inventory could be helpful in educating and following up patients undergoing peritoneal dialysis.

Aichi cohort study of the prognosis in patients newly initiated into dialysis (AICOPP): baseline characteristics and trends observed in diabetic nephropathy.

BACKGROUND: Over 300,000 patients receive maintenance dialysis in Japan; managing these patients is extremely important. This study aimed to report on prior management of chronic kidney diseases and prognostication after dialysis initiation. PATIENTS AND METHODS: Seventeen institutions participated in the Aichi cohort study of prognosis in patients newly initiated into dialysis and recruited patients over a period of 2 years. Exclusion criteria were (1) patients under 20 years; (2) patients who died before hospital discharge; and (3) patients who could not provide consent. RESULT: Here, we showed data on dialysis initiation time. Of 1524 patients with mean age of 67.5 ± 13.0 years, 659 patients were put on dialysis following diabetic nephropathy diagnosis. At dialysis initiation time, creatinine and estimated glomerular filtration rate levels were 8.97 ± 3.21 mg/dl and 5.45 ± 2.22 ml/min/1.73 m2, respectively. Medications taken were angiotensin II receptor blockers in 866; angiotensin-converting enzyme inhibitors in 135; calcium antagonist in 1202; and diuretics, alone or in combination, in 1059. Among patients with diabetic nephropathy, many had increased body weight and systolic blood pressure and were taking loop and thiazide diuretics at dialysis initiation time. Many patients with diabetic nephropathy had coronary artery disease and percutaneous coronary intervention. CONCLUSION: Many patients with diabetic nephropathy who registered for this study had coronary artery disease and problems with excess body fluid. Further analyses may clarify how underlying conditions and disease management before and after dialysis initiation affect prognosis.

Dynamics of Centrocestus armatus Transmission in Endemic River in Hyogo Prefecture, Japan.

Centrocestus armatus is an intestinal parasite belonging to the family Heterophyidae. We developed an apparatus for recovering cercariae and clarified the infection dynamics of this parasite. To clarify the circadian rhythm of cercarial shedding in the summer season, we filtrated 30 l of river water every 2 h for 24 h. Cercariae were first detected between 06:00 and 08:00 h, increased over time to reach peak at 16:00 h and decreased thereafter, thus showing a single-peak pattern. In a survey of seasonal change, approximately 200 cercariae were contained in 1 l of river water during the summer season, while none were found during the winter. This cercarial shedding pattern appeared to be related to sunrise/sunset and water/atmosphere temperature. Therefore, we examined whether cercarial shedding was affected by light or temperature changes under laboratory conditions, and confirmed that both light and temperature were important factors for cercarial shedding. Light was a stronger factor than water temperature. Cercarial shedding of C. armatus occurred in response to temperature and light. The change in the number of juvenile metacercariae detected in fish brain corresponded with monthly detection rates of cercariae; however, the incidence of new infections decreased in August. This suggests that Nipponocypris temminkii contains a defense mechanism against new infections that may have hindered the increase in parasite infectivity. These results clarified the smooth infection from the first to the second intermediate host of C. armatus in the endemic river. Throughout the study period, fecal samples were collected from 19 kites, 114 herons, and three unidentified species. However, our results using C. armatus showed a low value of 1% in herons and 5% in kites. The infection dynamics of final host to first intermediate host need to be further investigated.

Embryonic development of a whirligig beetle, Dineutus mellyi, with special reference to external morphology (insecta: Coleoptera, Gyrinidae).

The egg morphology and successive changes of developing embryos of the whirligig beetle, Dineutus mellyi (Adephaga: Gyrinidae) are described from observations based on light and scanning electron microscopy. The egg surface is characterized by minute conical projections covering the entire egg surface, a stalk-like micropylar projection at the anterior pole of the egg, and a longitudinal split line along which the chorion is cleaved during the middle embryonic stages. The germ band or embryo is formed on the ventral egg surface, and develops on the surface throughout the egg period; thus, the egg is a superficial type, as is the case in most coleopteran species. A pair of lateral tracheal gills (LTGs) of the first abdominal segment originates from appendage-like projections arising at the lateral side of pleuropodia, and the LTGs of the second to ninth abdominal segments are arranged in a row with that of the first segment. Therefore, LTGs are structures with serial homology. The paired dorsal tracheal gills (DTGs) of the ninth abdominal segment are formed on the regions just latero-dorsal to the LTGs of this segment. Regarding the pleuropodia as the structures being homologous with thoracic legs, neither the LTGs nor DTGs are homologous with thoracic legs, but originate in the more lateral region corresponding to the future pleura of the thoracic segments. The last (10th) abdominal segment in the larva is formed by the fusion of the embryonic 10th and 11th abdominal segments. Four terminal hooks at the end of the last abdominal segment originate from two pairs of swellings on the posterior end of the embryonic 11th abdominal segment. It is proposed that the terminal hooks possibly correspond to the claws of medially fused cerci of the embryonic 11th abdominal segment.