Plasma cell infiltration and treatment effect in breast cancer patients treated with neoadjuvant chemotherapy.

BACKGROUND: Tumour-infiltrating lymphocyte (TIL)-high breast tumours have a high rate of pathological complete response (pCR) with neoadjuvant chemotherapy. In our routine pathological diagnoses of biopsy specimens from pCR cases, we have observed a high infiltration of plasma cells (PCs). A positive correlation of PCs with favourable patient outcome has recently been reported, but little is known about how PCs contribute to local tumour immunity. METHODS: We retrospectively examined biopsy specimens from 146 patients with invasive breast cancer who received neoadjuvant chemotherapy. CD138+ PC infiltration was assessed by immunohistochemistry. Multiplexed fluorescent immunohistochemistry (mfIHC) with T and B cell markers was also conducted to elucidate the profile of immune cells. RESULTS: Greater PC infiltration was observed in the pCR group (p = 0.028) and this trend was confirmed in another patient cohort. With mfIHC, we observed significantly more CD8+, T-bet+CD4+, and CD8+FOXP3+ T cells, total B cells and PCs in pCR cases. Such cases were also characterised by high expression of both PD-1 and PD-L1 on B cells and PCs. In patients with hormone receptor-negative tumours, high PC infiltration was correlated with significantly longer disease-free survival (p = 0.034). CONCLUSIONS: We found that higher PC infiltration in biopsy specimens before neoadjuvant chemotherapy was associated with pCR. With mfIHC, we also revealed that the local cytotoxic immune response was clearly enhanced in pCR cases, as was the infiltration of B cells including PCs. Moreover, higher PC levels were correlated with favourable outcomes in hormone receptor-negative breast cancer patients.

Hyaluronan synthesis supports glutamate transporter activity.

Hyaluronan is synthesized, secreted, and anchored by hyaluronan synthases (HAS) at the plasma membrane and comprises the backbone of perineuronal nets around neuronal soma and dendrites. However, the molecular targets of hyaluronan to regulate synaptic transmission in the central nervous system have not been fully identified. Here, we report that hyaluronan is a negative regulator of excitatory signals. At excitatory synapses, glutamate is removed by glutamate transporters to turn off the signal and prevent excitotoxicity. Hyaluronan synthesized by HAS supports the activity of glial glutamate transporter 1 (GLT1). GLT1 also retracted from cellular processes of cultured astrocytes after hyaluronidase treatment and hyaluronan synthesis inhibition. A serial knockout study showed that all three HAS subtypes recruit GLT1 to cellular processes. Furthermore, hyaluronidase treatment activated neurons in a dissociated rat hippocampal culture and caused neuronal damage due to excitotoxicity. Our findings reveal that hyaluronan helps to turn off excitatory signals by supporting glutamate clearance. Cover Image for this issue: doi: 10.1111/jnc.14516.

Analysis of circulating tumour cell and the epithelial mesenchymal transition (EMT) status during eribulin-based treatment in 22 patients with metastatic breast cancer: a pilot study.

BACKGROUND: Liquid biopsy approaches, such as measuring circulating tumour cells (CTCs), have recently been introduced in several clinical studies. However, the development of CTCs as a predictive marker for treatment effects on breast cancer remains an enormous task. We investigated CTCs, including epithelial mesenchymal transition (EMT) status, from metastatic breast cancer patients who had received eribulin-based treatment, which reportedly suppresses EMT as a means of tumour suppression. Our aim was to test the possibility of this method serving as a tool predicting eribulin efficacy. METHODS: Twenty-two patients were enrolled and peripheral blood samples were collected before eribulin treatment. CTCs were then examined using a Microfluidic Chip device. CTCs positive for vimentin and pan-cytokeratin were defined as mesenchymal and epithelial CTCs, respectively. Progression-free survival (PFS) and clinical response were assessable in 20 and 18 patients, respectively, in relation to the number of CTCs. RESULTS: Numbers of total CTCs were significantly increased in patients with progressive disease during treatment (p = 0.006). Median PFS was 14.6 weeks and patients with more total and mesenchymal CTCs at baseline had significantly shorter PFS (p = 0.0013 and 0.013, respectively). Multivariate logistic regression analysis revealed small number of total baseline CTCs and long disease-free survival to be related to long PFS (p = 0.0004 and 0.020, respectively). CONCLUSIONS: Our data suggest that determining both mesenchymal and epithelial CTCs at baseline might be a good tool for predicting eribulin responsiveness. Evaluation of mesenchymal CTC can be considered as a parameter in larger studies, while most clinical trials are currently employing only the detection of the epithelial cellular adhesion molecule (EpCAM).

Tissue Reaction to a Novel Bone Substitute Material Fabricated With Biodegradable Polymer-Calcium Phosphate Nanoparticle Composite.

PURPOSE: The aim of this study was to evaluate the effectiveness of a novel bone substitute material fabricated using a biodegradable polymer-calcium phosphate nanoparticle composite. METHODS: Porous structured poly-L-lactic acid (PLLA) and hydroxyapatite (HA) nanoparticle composite, which was fabricated using solid-liquid phase separation and freeze-drying methods, was grafted into bone defects created in rat calvarium or tibia. Rats were killed 4 weeks after surgery, and histological analyses were performed to evaluate new bone formation. RESULTS: Scanning electron microscopic observation showed the interconnecting pores within the material and the pore diameter was approximately 100 to 300 µm. HA nanoparticles were observed to be embedded into the PLLA beams. In the calvarial implantation model, abundant blood vessels and fibroblastic cells were observed penetrating into pores, and in the tibia model, newly formed bone was present around and within the composite. CONCLUSIONS: The PLLA-HA nanoparticle composite bone substitute developed in this study showed biocompatibility, elasticity, and operability and thus has potential as a novel bone substitute.

Ki67 expression and the effect of neo-adjuvant chemotherapy on luminal HER2-negative breast cancer.

BACKGROUND: Patients with luminal HER2-negative tumours have a favourable prognosis. However, there is a subpopulation in which poorer outcomes are obtained with endocrine therapy alone. This subpopulation is considered to benefit from chemotherapy. However, the significance of chemotherapy for those with luminal tumours has decreased due to recent changes in treatment strategies. Thus, it is often difficult to determine whether we should recommend chemotherapy to such patients in clinical practice. We investigated Ki67 expression, as a means of predicting the responses of luminal HER2-negative breast cancer patients to neo-adjuvant chemotherapy (NAC), in order to identify a subpopulation that would benefit from these treatments. METHODS: We enrolled 114 luminal HER2-negative breast cancer patients undergoing surgery after NAC. Biomarkers were examined using biopsy specimens obtained prior to treatment, to avoid any chemotherapy-related effects. Chemotherapy effects were determined employing operative specimens and we defined pathological complete response (pCR) as invasive nest disappearance, based only on the primary breast tumour. We applied receiver operating characteristic curve analysis to data from our 114 patients, to investigate Ki67 expression as a predictor of pCR. RESULTS: The pCR rate was significantly higher for tumours with high Ki67 expression (p < 0.01) and all patients who obtained pCR remained recurrence-free during the median 58-month observation period. We identified 35% as the Ki67 cut-off value which distinguishes those with a pCR from other cases. Another dataset, comprised of 196 patients with a median 29-month observation period, was recruited for validation. Disease-free survival was found to be significantly (p < 0.01) lower in the patients with tumours in which Ki67 expression was higher than 35%. CONCLUSION: Our results raise the possibility of the luminal HER2-negative subpopulation with Ki67 expression higher than 35% benefiting from chemotherapy, as evidenced by improved survival.

Cross-talk of receptor activator of nuclear factor-κB ligand signaling with renin-angiotensin system in vascular calcification.

OBJECTIVE: Vascular calcification is accelerated by hypertension and also contributes to hypertension; however, it is an enigma why hypertension and vascular calcification are a vicious spiral. The present study elucidates the cross-talk between renin-angiotensin II system and receptor activator of nuclear factor-κB ligand (RANKL) system in vascular calcification. APPROACH AND RESULTS: Angiotensin (Ang) II (10(-7) mol/L) significantly increased calcium deposition as assessed by Alizarin Red staining, associated with a significant increase in the expression of RANKL, RANK, and bone-related genes, such as cbfa1 and msx2, in human aortic vascular smooth muscle cells. Infusion of Ang II (100 ng/kg per minute) in ovariectomized ApoE(-/-) mice under high-fat diet significantly increased the expression of RANKL system and calcification in vivo, whereas administration of Ang II receptor blocker (olmesartan, 3 mg/kg per day) decreased the calcification and bone markers' expression. In addition, male OPG(-/-) mice showed a significant increase in vascular calcification followed by Ang II infusion as compared with wild type. Conversely, RANKL significantly increased Ang II type 1 receptor and angiotensin II-converting enzyme expression in vascular smooth muscle cells via extracellular signal-regulated protein kinase phosphorylation. CONCLUSIONS: The present study demonstrated that Ang II significantly induced vascular calcification in vitro and in vivo through RANKL activation. In addition, RANKL activated renin-angiotensin II system, especially angiotensin II-converting enzyme and Ang II type 1 receptor. Cross-talk between renin-angiotensin II system and RANKL system might work as a vicious cycle to promote vascular calcification in atherosclerosis. Further studies to inhibit renin-angiotensin II system and RANKL may provide new therapeutic options to prevent and regress vascular calcification.

Nanosized zinc oxide particles do not promote DHPN-induced lung carcinogenesis but cause reversible epithelial hyperplasia of terminal bronchioles.

Zinc oxide (ZnO) is known to induce lung toxicity, including terminal bronchiolar epithelial hyperplasia, which gives rise to concerns that nanosized ZnO (nZnO) might lead to lung carcinogenesis. We studied the tumor promoting activity of nZnO by an initiation-promotion protocol using human c-Ha-ras proto-oncogene transgenic rats (Hras128 rats). The rats were given 0.2 % N-nitrosobis(2-hydroxypropyl)amine (DHPN) in the drinking water for 2 weeks and then treated with 0.5 ml of 250 or 500 µg/ml nZnO suspension by intra-pulmonary spraying once every 2 weeks for a total of 7 times. Treatment with nZnO particles did not promote DHPN-induced lung carcinogenesis. However, nZnO dose-dependently caused epithelial hyperplasia of terminal bronchioles (EHTB) and fibrosis-associated interstitial pneumonitis (FAIP) that were independent of DHPN treatment. Tracing the fate of EHTB lesions in wild-type rats indicated that the hyperplastic lesions almost completely disappeared within 12 weeks after the last nZnO treatment. Since nZnO particles were not found in the lung and ZnCl2 solution induced similar lung lesions and gene expression profiles, the observed lesions were most likely caused by dissolved Zn(2+). In summary, nZnO did not promote carcinogenesis in the lung and induced EHTB and FAIP lesions that regressed rapidly, probably due to clearance of surplus Zn(2+) from the lung.

[Bone metabolism and cardiovascular function update. Cross link of hypertension, bone loss and vascular calcification - common back grounds in renin angiotensin system with anti-aging aspect -].

Hypertension, osteoporosis and vascular calcification are major diseases in the recent aging society and may share the same backgrounds genetically and environmentally. As treatments to prevent aging-related diseases simultaneously are desirable, we investigate common backgrounds underling these diseases. Renin- angiotensin system, which causes high blood pressure, is found to be involved in bone metabolism. Angiotensin II has been shown to accelerate osteoporosis through RANKL up-regulation in osteoblast. RANKL, in turn, contributes to vascular calcification by regulating bone morphogenetic protein-2 and MGP expression, as well as bone-related proteins. Angiotensin type 1 receptor blockers (ARBs) ameliorate osteoporosis and vascular calcification beyond their blood pressure lowering effects. These pleiotropic effects of antihypertensive drugs such as ARBs might benefit especially hypertensive postmenopausal women.

Exploration of reference genes for the development of a diagnostic kit on vascular aging in human saliva.

Identifying reliable biomarkers in saliva can be a promising approach to developing a rapid diagnostic kit for detecting vascular aging. This study investigated the most suitable reference gene for polymerase chain reaction (PCR) in saliva that is not affected by vascular aging variables. Whole saliva samples were collected to assess the expression of reference genes: actin beta (ACTB), 18S ribosomal RNA (18S rRNA), beta-2-microglobulin, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The most abundantly expressed gene was 18S rRNA, and the least expressed gene was GAPDH. Four genes were ranked according to their relative stability, as determined by mathematical algorithms, indicating that ACTB and 18S rRNA were stably expressed as reference genes. 18S rRNA was identified as the most promising reference gene for detecting systemic diseases using saliva from patients with vascular aging in these limited experimental conditions.

Increase in antibody titer and change over time associated with severe acute respiratory syndrome coronavirus 2 infection after mRNA vaccination: Consideration of the significance of additional vaccination.

Key Clinical Message: Most Japanese patients naturally infected with COVID-19 were infected after mRNA vaccination, and many maintained high antibody titers due to hybrid immunity. The significance of additional vaccination in hybrid-immunized cases is highly questionable. Abstract: Spontaneous infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) after mRNA vaccination causes a marked increase in antibody titer because of the combined effect of vaccine and infection ("hybrid immunity"). In this study, we discuss the significance of the mRNA vaccine booster inoculation that has been repeatedly performed in Japan. We describe the temporal trends of antibody titers in cases in which antibody titers were markedly increased by hybrid immunization. The antibody titer increased with hybrid immunization and tended to decrease with time. However, several cases maintained high antibody titers for approximately 1 year after coronavirus disease 2019 (COVID-19) diagnosis, even without booster vaccination. Most Japanese patients naturally infected with COVID-19 were infected after mRNA vaccination, and many maintained high antibody titers due to hybrid immunity. The significance of additional vaccination in hybrid-immunized cases is highly questionable regarding cost-effectiveness and risk-benefit.

FMRP, FXR1 protein and Dlg4 mRNA, which are associated with fragile X syndrome, are involved in the ubiquitin-proteasome system.

The ubiquitin-proteasome system (UPS) is a proteolytic pathway that is essential for life maintenance and vital functions, and its disruption causes serious impairments, e.g., disease development. Thus, the UPS is properly regulated. Here we show novel UPS-related factors: the fragile X mental retardation 1 (FMR1) and Fmr1 autosomal homolog 1 (FXR1) proteins and discs large MAGUK scaffold protein 4 (Dlg4) mRNA, which are associated with Fragile X syndrome, are involved in UPS activity. Fmr1-, Fxr1- and Dlg4-knockdown and Fmr1- and Fxr1-knockdown resulted in increased ubiquitination and proteasome activity, respectively. FXR1 protein was further confirmed to be associated with proteasomes, and Dlg4 mRNA itself was found to be involved in the UPS. Knockdown of these genes also affected neurite outgrowth. These findings provide new insights into the regulatory mechanism of the UPS and into the interpretation of the pathogenesis of diseases in which these genes are involved as UPS-related factors.

Association between oral frailty and cystatin C-related indices-A questionnaire (OFI-8) study in general internal medicine practice.

BACKGROUND: Cystatin C-related indices such as the ratio of creatinine to cystatin C (Cr/CysC) and the ratio of estimated glomerular filtration rate by cystatin C (eGFRcys) to creatinine eGFRcre (eGFRcys/eGFRcre) levels have been shown to be associated with muscle mass and strength and can be markers of sarcopenia. Oral frailty is defined as an age-related gradual loss of oral functions, accompanied by a decline in cognitive and physical functions. It results in adverse health-related outcomes in older age, including mortality, physical frailty, functional disability, poor quality of life, and increased hospitalization and falls. Therefore, poor oral health among the elderly is an important health concern due to its association with the pathogenesis of systemic frailty, suggesting it to be a multidimensional geriatric syndrome. The Oral Frailty Index-8 (OFI-8) is a questionnaire that can be used for easy screening of oral frailty. This study aimed to investigate whether cystatin C- related indices are different between patients with low to moderate risk of oral frailty and those at high risk of oral frailty, using the OFI-8 in attending a general internal medicine outpatient clinic. MATERIALS AND METHODS: This is a cross-sectional study that included 251 patients with a mean age of 77.7±6.6 years and a median age of 77 years (128 men: mean age, 77.1±7.3 years; median age, 77 years and 123 women: mean age, 78.4±5.7 years; median age, 78 years) attending general internal medicine outpatient clinics. OFI-8 scores were tabulated by gender to determine whether there were differences between patients at low to moderate risk of oral frailty (OFI-8 score ≤3 points) and those at high risk (OFI-8 score ≥4 points) in Cr/CysC, eGFRcys/eGFRcre levels, height, weight, grip strength, etc. were examined. RESULTS: The OFI-8 score was higher in women than in men, suggesting that oral frailty is more common in women. Cr/CysC, eGFRcys/eGFRcre and grip strength were significantly lower in both men and women in the high-risk group for oral frailty (OFI-8 score ≥ 4). Height, hemoglobin level, red blood cell count, and serum albumin levels were significantly lower in men with an OFI-8 score ≥4. Receiver operating characteristic curve (ROC) analysis also showed that Cr/CysC and eGFRcys/eGFRcre were significantly associated with an OFI-8 score≥4 in both men and women. CONCLUSION: Cr/CysC and eGFRcys/eGFRcre were significantly lower in the high-risk group for oral frailty on the OFI-8in both men and women. A relationship exists among cystatin C-related indices, which can effectively screen systemic frailty. Similarly, the OFI-8 score can be used to effectively screen oral frailty. Thus, a collaboration that incorporates both systemic and oral frailty from medical and dental perspectives is required.

Trends in Antibody Titers after SARS-CoV-2 Vaccination-Insights from Self-Paid Tests at a General Internal Medicine Clinic.

Background: The rise in antibody titers against the novel coronavirus (SARS-CoV-2) and its duration are considered an important indicator for confirming the effect of a COVID-19 vaccine, and self-paid tests of antibody titer are conducted in many facilities nationwide. Methods: The relationship between the number of days after the second and third dose of vaccines, age, and antibody titer was determined from the medical records of general internal medicine clinics that conducted self-paid testing of the SARS-CoV-2 antibody titer using Elecsys Anti-SARS-CoV-2 S (Roche Diagnostics); the relationship between the number of days after two or more doses of vaccines and antibody titer was also determined. We also examined the antibody titers in cases of spontaneous infection with SARS-CoV-2 after two or more doses of the vaccine. Results: Log-transformed SARS-CoV-2 antibody titers measured within 1 month from the second or third dose of vaccine showed a negative correlation with age (p < 0.05). In addition, the log-transformed antibody titers also showed a negative correlation trend with the number of days after the second dose of vaccine (p = 0.055); however, there were no significant correlations between the log-transformed antibody titers and the number of days after the third dose of vaccine. The median antibody titer after the third vaccination was 18,300 U/mL, more than 10 times the median antibody titer after the second dose of vaccine, of 1185 U/mL. There were also some cases of infection after the third or fourth dose of vaccine, with antibody titers in the tens of thousands of U/ml after infection, but the patients still received further booster vaccinations after the infection. Conclusions: The antibody titers after the third vaccination did not attenuate after a short follow-up period of one month, while they tended to attenuate after the second vaccination. It is considered that many people in Japan received further booster vaccinations after spontaneous infection, even though they already had antibody titers in the tens of thousands of U/mL due to "hybrid immunity" after spontaneous infection following two or more doses of vaccine. The clinical significance of the booster vaccination in this population still needs to be thoroughly investigated and should be prioritized for those with low SARS-CoV-2 antibody titers.

Longitudinal Changes in IgG-Type SARS-CoV-2 Antibody Titers after COVID-19 Vaccination and a Prominent Increase in Antibody Titers When Infected after Vaccination.

OBJECTIVE: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody titers level and duration of elevated levels are considered important indicators for confirming the efficacy of coronavirus disease 2019 (COVID-19) vaccines. The objective of this study was to demonstrate the changes in antibody titers after the second and third doses of the COVID-19 vaccine, and to determine the antibody titers in cases of spontaneous infection with SARS-CoV-2 after vaccination. MATERIALS AND METHODS: From June 2021 to February 2023, IgG-type SARS-CoV-2 antibody titers were measured in 127 participants, including 74 outpatients and 53 members of staff, at the Osaka Dental University Hospital (64 males and 63 females, mean age 52.3 ± 19.0 years). RESULTS: Consistent with previous reports, the SARS-CoV-2 antibody titer decreased with time, not only after the second dose but also after the third dose of the vaccine if there was no spontaneous COVID-19 infection. We also confirmed that the third booster vaccination was effective in increasing the antibody titer. A total of 21 cases of natural infections were observed after administering two or more doses of the vaccine. Thirteen of these patients had post-infection antibody titers exceeding 40,000 AU/mL, and some cases continued to maintain antibody titers in the tens of thousands of AU/mL even after more than 6 months had passed since infection. CONCLUSIONS: The rise in and duration of antibody titers against SARS-CoV-2 are considered important indicators for confirming the efficacy of novel COVID-19 vaccines. A longitudinal follow-up of antibody titers after vaccination in larger studies is warranted.

Multi-walled carbon nanotubes translocate into the pleural cavity and induce visceral mesothelial proliferation in rats.

Multi-walled carbon nanotubes have a fibrous structure similar to asbestos and induce mesothelioma when injected into the peritoneal cavity. In the present study, we investigated whether carbon nanotubes administered into the lung through the trachea induce mesothelial lesions. Male F344 rats were treated with 0.5 mL of 500 µg/mL suspensions of multi-walled carbon nanotubes or crocidolite five times over a 9-day period by intrapulmonary spraying. Pleural cavity lavage fluid, lung and chest wall were then collected. Multi-walled carbon nanotubes and crocidolite were found mainly in alveolar macrophages and mediastinal lymph nodes. Importantly, the fibers were also found in the cell pellets of the pleural cavity lavage, mostly in macrophages. Both multi-walled carbon nanotube and crocidolite treatment induced hyperplastic proliferative lesions of the visceral mesothelium, with their proliferating cell nuclear antigen indices approximately 10-fold that of the vehicle control. The hyperplastic lesions were associated with inflammatory cell infiltration and inflammation-induced fibrotic lesions of the pleural tissues. The fibers were not found in the mesothelial proliferative lesions themselves. In the pleural cavity, abundant inflammatory cell infiltration, mainly composed of macrophages, was observed. Conditioned cell culture media of macrophages treated with multi-walled carbon nanotubes and crocidolite and the supernatants of pleural cavity lavage fluid from the dosed rats increased mesothelial cell proliferation in vitro, suggesting that mesothelial proliferative lesions were induced by inflammatory events in the lung and pleural cavity and likely mediated by macrophages. In conclusion, intrapulmonary administration of multi-walled carbon nanotubes, like asbestos, induced mesothelial proliferation potentially associated with mesothelioma development.

Estrogen inhibits vascular calcification via vascular RANKL system: common mechanism of osteoporosis and vascular calcification.

RATIONALE: Arterial calcification and osteoporosis are associated in postmenopausal women. RANK (the receptor activator of nuclear factor kappaB), RANKL (RANK ligand), and osteoprotegerin are key proteins in bone metabolism and have been found at the site of aortic calcification. The role of these proteins in vasculature, as well as the contribution of estrogen to vascular calcification, is poorly understood. OBJECTIVE: To clarify the mechanism of RANKL system to vascular calcification in the context of estrogen deficiency. METHODS AND RESULTS: RANKL induced the calcification inducer bone morphogenetic protein-2 by human aortic endothelial cells (HAECs) and decreased the calcification inhibitor matrix Gla protein (MGP) in human aortic smooth muscle cells (HASMCs), as quantified by real-time PCR and Western blot analysis. RANKL also induced bone-related gene mRNA expression and calcium deposition (Alizarin red staining) followed by the osteogenic differentiation of HASMCs. Estrogen inhibited RANKL signaling in HAECs and HASMCs mainly through estrogen receptor alpha. Apolipoprotein E-deficient mice fed with Western high-fat diet for 3 months presented atherosclerotic calcification (Oil red and Alizarin red staining) and osteoporosis (microcomputed tomographic analysis) after ovariectomy and increased expression of RANKL, RANK, and osteopontin in atherosclerotic lesion, as detected by in situ hybridization. Estrogen replacement inhibited osteoporosis and the bone morphogenetic protein osteogenic pathway in aorta by decreasing phosphorylation of smad-1/5/8 and increasing MGP mRNA expression. CONCLUSIONS: RANKL contributes to vascular calcification by regulating bone morphogenetic protein-2 and MGP expression, as well as bone-related proteins, and is counteracted by estrogen in a receptor-dependent manner.

Role of osteopontin in early phase of renal crystal formation: immunohistochemical and microstructural comparisons with osteopontin knock-out mice.

Osteopontin (OPN) is an important matrix protein of renal calcium stone. However, the function of OPN in the early phase of renal crystal formation is not well defined. In this study, we examined OPN expression in the early phase of renal crystal formation with ultra-microstructural observations and immuno-TEM (transmission electron microscopy) in control and OPN knock-out (OPN-KO) mice. Glyoxylate (100 mg/kg) was intra-abdominally administered to male wild-type mice (C57BL/6, 8 weeks of age) and OPN-KO mice (C57BL/6, 8 weeks of age). Kidney was collected before and 6, 12, and 24 h after administration. We examined the relation between renal crystal formation and microstructural OPN location using TEM and immunohistochemical staining of OPN as well as western blotting and quantitative RT-PCR for OPN. OPN protein expression gradually increased in the renal cortex-medulla junction after glyoxylate administration, and OPN mRNA was increased until 12 h, but decreased at 24 h. In ultra-microstructural observation, OPN began to appear on the luminal side of renal distal tubular cells at 6 h and was gradually detected in the tubular lumen at 12 h. OPN was present in the crystal nuclei and collapsed mitochondria in the tubular lumen. In the OPN-KO mice, collapsed mitochondria were present, but no crystal nuclei formation were detected at 24 h. Based on the results this study proposed that the appearance of organelles, such as mitochondria and microvilli, in the tubular lumen after cell injury may be the starting point of crystal nucleus formation due to the aggregation ability of OPN.

[Activation of osteoclasts by RAAS and strategy of target therapy on bone metabolic diseases].

Angiotensin II is a potent stimulator of vascular smooth mustle cells that increases blood pressure, and angiotensin type 1 receptors have been identified on osteoblasts; thus, the renin-angiotensin system has been suggested to be involved in bone metabolism. The authors reported that angiotensin II significantly increased TRAP-positive multinuclear osteoclasts with the up-regulation of RANKL expression through extracellular kinase of osteoblast. These effects were abolished with co-treatment of angiotensin converting enzyme (ACE) inhibitors or angiotensin type 1 receptor blockers (ARB). Recently, RANK-RANKL-OPG system is proved to be involved in vascular calcification and RAAS also might be involved in the development of the disease. Thus, target therapy for RAAS might bring additional beneficial effects on bone metabolic diseases such as osteoporosis, rheumatoid arthritis and vascular calcification other than blood pressure lowering effect.

[Study on the Concentration of Radioactive Cesium in the Environment after the Fukushima Daiichi Nuclear Power Plant Accident].

The Tohoku-Pacific Ocean Earthquake that occurred on March 11, 2011 and the resulting tsunami caused the loss of many people and extensive damage in a wide area. Among the anthropogenic radionuclides dispersed from the Fukushima Daiichi Nuclear Power Plant, 134Cs and 137Cs have very long half-lives of approximately 2 years and 30 years, respectively, and there are concerns about their uptake into soil and living things. This paper describes a study conducted by the authors' group on radiocesium activity concentrations in the environment.

DISTRIBUTION OF GAMMA RADIATION DOSE RATE AND ACTIVITY CONCENTRATION IN SOIL RELATED WITH NATURAL RADIONUCLIDES ON TAIWAN MAIN ISLAND.

To enable precise assessment of health impacts following a nuclear power plant accident, extensive and detailed data on environmental radiation levels are needed. This study was undertaken to investigate the air and the soil radiation levels using a car-borne survey on the main island of Taiwan where no extensive environmental radiation distribution survey had been conducted before. The mean air absorbed dose rate on this island was 57 ± 10 nGy h-1. The measured dose rate distribution varied depending on the geology of the soils, and ranged from 22 to 113 nGy h-1. The mean radiation level in soil was 539 ± 124 Bq kg-1 for 40K, 23 ± 8 Bq kg-1 for 238U and 41 ± 22 Bq kg-1 for 232Th. The air absorbed dose rate (58 nGy h-1) calculated from these data of mean radiation level in soil was comparable to that determined by the car-borne survey method. Thus, this study yielded detailed data on air absorbed dose rate depending primarily on the geology of the soils on the main island of Taiwan.