Clinicopathological risk factors for a poor prognosis of primary central nervous system lymphoma in elderly patients in the Tohoku and Niigata area: a multicenter, retrospective, cohort study of the Tohoku Brain Tumor Study Group.

Clinicopathological risk factors for a poor prognosis were investigated in elderly patients with malignant lymphoma of the central nervous system. A total of 82 pathologically confirmed, CD20-positive, diffuse large B-cell lymphoma patients aged 71 years or older who underwent therapeutic intervention in the Tohoku and Niigata area in Japan were retrospectively reviewed. A univariate analysis was performed by the log-rank test using the Kaplan-Meier method. A Cox proportional hazards model was used for multivariate analysis of risk factors. Of the 82 patients, 39 were male and 43 were female, and their median age at onset was 75 years. At the end of the study, there were 34 relapse-free patients (41.5%), 48 relapse cases (58.5%), median progression-free survival was 18 months, and median overall survival (OS) was 26 months; there were 41 deaths and 41 survivors. Multivariate analysis of median OS showed that Karnofsky Performance Status less than 60% 3 months after treatment (p = 0.022, hazard ratio (HR) = 2.591) was the clinical risk factor, and double expressor lymphoma (p = 0.004, HR = 3.163), expression of programmed death-ligand 1 in tumor infiltrating lymphocytes or tumor-associated macrophages (p < 0.001, HR = 5.455), and Epstein-Barr virus infection (p = 0.031, HR = 5.304) were the pathological risk factors.

Oxidative Stress Mediates Vascular Tortuosity.

Vascular tortuosity is associated with various disorders and is being increasingly detected through advances in imaging techniques. The underlying mechanisms for vascular tortuosity, however, remain unclear. Here, we tested the hypothesis that oxidative stress mediates the generation of tortuous vessels. We used the bilateral common carotid artery (CCA) ligation model to induce vascular tortuosity. Both young and adult rats showed basilar artery tortuous morphological changes one month after bilateral CCA ligation. These tortuous changes were permanent but more pronounced in the adult rats. Microarray and real-time PCR analysis revealed that these tortuous changes were accompanied by the induction of oxidative stress-related genes. Moreover, the indicated model in rabbits showed that tortuous morphological changes to the basilar artery were suppressed by antioxidant treatment. These results are highly suggestive of the significance of oxidative stress in the development of vascular tortuosity. Although further studies will be needed to elucidate the possible mechanisms by which oxidative stress enhances vascular tortuosity, our study also points toward possible prophylaxis and treatment for vascular tortuosity.

Combination chemoradiotherapy with temozolomide, vincristine, and interferon-ß might improve outcomes regardless of O6-methyl-guanine-DNA-methyltransferase (MGMT) promoter methylation status in newly glioblastoma.

BACKGROUND: This investigator-initiated, open-label, single-arm, single-institute study was conducted to investigate the effectiveness of induction combination chemoradiotherapy and long-term maintenance therapy with temozolomide (TMZ) plus interferon (IFN)-ß for glioblastoma. METHODS: The initial induction combination chemoradiotherapy comprised radiotherapy plus TMZ plus vincristine plus IFN-ß. Maintenance chemotherapy comprised monthly TMZ, continued for 24-50 cycles, plus weekly IFN-ß continued for as long as possible. The primary endpoint was 2-year overall survival (2y-OS). The study protocol was to be considered valid if the expected 2y-OS was over 38% and the lower limit of the 95% confidence interval (CI) was no less than 31.7% compared with historical controls, using Kaplan-Meier methods. Secondary endpoints were median progression-free survival (mPFS), median OS (mOS), 5-year OS rate (5y-OS), and mPFS and mOS classified according to MGMT promoter methylation status. RESULTS: Forty-seven patients were analyzed. The 2y-OS was 40.7% (95%CI, 27.5-55.4%). The mPFS and mOS were 11.0 months and 18.0 months, respectively, and 5y-OS was 20.3% (95%CI, 10.9-34.6%). The mPFS in groups with and without MGMT promoter methylation in the tumor was 10.0 months and 11.0 months (p = 0.59), respectively, and mOS was 24.0 months and 18.0 months (p = 0.88), respectively. The frequency of grade 3/4 neutropenia was 19.1%. CONCLUSIONS: The 2y-OS with induction multidrug combination chemoradiotherapy and long-term maintenance therapy comprising TMZ plus IFN-ß tended to exceed that of historical controls, but the lower limit of the 95%CI was below 31.7%. Although the number of cases was small, this protocol may rule out MGMT promoter methylation status as a prognostic factor. TRIAL REGISTRATION: University Hospital Medical Information Network (number UMIN000040599 ).

Irreversible Neuronal Damage Begins Just After Aneurysm Rupture in Poor-Grade Subarachnoid Hemorrhage Patients.

Pathophysiological findings of early brain injury in humans have not permitted conclusive determinations. We explored the essence of this phenomenon by taking intraoperative cortical specimens of Hunt-Kosnik grades IV~V (poor-grade) subarachnoid hemorrhages (SAH). From 2013 to 2017, we treated 39 consecutive poor-grade patients in 226 cases of aneurysmal SAH. Fourteen of the 39 patients agreed to this study following written informed consent. We took specimens from untouched areas prior to surgical intervention: cortex near the ruptured aneurysm for clipping, convexity cortex for cerebral ventricular drainage. Cortical specimens were stained with hematoxylin-eosin, anti-cleaved caspase-3, and anti-DNA/RNA damage staining. Positive signals were calculated in six random, high-power fields for quantitative assessment. Double immunofluorescence was done to evaluate neural damage. Chi-square analyses were carried out to assess the correlation between the Glasgow Outcome Scale at 90 days after the ictus and the number of positive cells. Cortical specimens were taken at 12.7 ± 7.00 h after the first ictus. All 14 cases showed dense nuclei, with the appearance of acidic and shrunken cytoplasms. Diffuse positivity of anti-cleaved caspase-3 and anti-DNA/RNA damage was detected. Cleaved caspase-3 was detected in 68% of neurons, and DNA/RNA damage was detected in 64% of neurons. Positive reactions of both antibodies indicated poor outcome. With poor-grade cases, irreversible ischemic, apoptotic, and oxidative changes were detected in the cerebral cortex within several hours after the ictus. Those changes occurred far from the aneurysm. Our findings suggest that a revolution is needed in the treatment strategy for poor-grade SAH.

Intra-arterial anti-oxidant power negatively correlates with white matter injury, and oxidative stress positively correlates with disability in daily activities.

Oxidative stress influences many kinds of diseases. Our hypothesis is that oxidative stress and antioxidant potentials correlate with cognitive function, activities of daily life and white matter injury. (UMIN-CTR R000016770) Thirty-two consecutive patients participated to this study after informed consent. A routine biochemical analysis, modified-Rankin Scale (m-RS), revised Hasegawa Dementia Scale (HDS-R), Mini-Mental State Examination (MMSE) and fluid-attenuated-inversion-recovery imaging (FLAIR) were performed before admission. Derivatives of reactive oxygen metabolites (d-ROMs) and biological antioxidant potential (BAP) were measured photometrically using arterial blood. Statistical analyses were done by analysis variance or logistic regression analysis. Median age was 72 (IQR: 64.3 -- 75.8). The d-ROMS were 367 ± 55.4, and BAP was 1967 ± 284. HDS-R and m-RS deteriorated with d-ROMs elevation (p < 0.05). Uric acid and creatinine decreased with d-ROMs elevation (p < 0.05). Both periventricular hyperintensity grade and deep and subcortical white matter hyperintensity grade worsened with BAP reduction (p < 0.05). Oxidative stress correlates negatively with cognitive function and activities of daily life. Low antioxidative potentials correlate with aggravation of white matter injury. We should control both oxidative stress and antioxidative potential to maintain healthy lives.

The Role of Intracranial Pressure and Subarachnoid Blood Clots in Early Brain Injury After Experimental Subarachnoid Hemorrhage in Rats.

OBJECTIVE: Early brain injury after subarachnoid hemorrhage (SAH), which is considered a main factor leading to poor outcome, is believed to be caused by the increase of intracranial pressure (ICP) and/or the presence of subarachnoid blood clots (SBC) itself. The purpose of this study was to examine whether ICP or SBC is more important to neurologic deficit in the presence of apoptosis or edema. METHODS: A total of 50 rats were allocated to 3 groups: an endovascular perforation SAH model (the SAH group), a cisterna magna saline injection model (the saline injection group), and a cisterna magna sham injection model (the sham injection group). Statistical analysis of correlations among the ICP, the grade of clot volume, neuronal apoptosis, brain water content (brain edema), and neurologic deficit was performed. RESULTS: In the SAH group, each of increased ICP and clot volume was correlated with neuronal apoptosis and brain edema. In the saline injection group, increased ICP was associated with apoptosis, but it did not correlate with brain edema. Neuronal apoptosis (r = 0.75; P < 0.01) and brain edema (r = 0.89; P < 0.01) correlated independently with neurologic deficit in the SAH group. CONCLUSIONS: The present study suggests that neuronal apoptosis is caused mainly by increased ICP, whereas brain edema is induced by SBC, and increased ICP could aggravate it in the presence of SBC. Brain edema could affect neurologic deficit, but apoptosis alone may be less influential. Not only ICP but also SBC seem important for brain damage in the acute stage of SAH.

The Role of Oxidative Stress in Microvascular Disturbances after Experimental Subarachnoid Hemorrhage.

Oxidative stress was shown to play a crucial role in the diverse pathogenesis of early brain injury (EBI) after subarachnoid hemorrhage (SAH). Microcirculatory dysfunction is thought to be an important and fundamental pathological change in EBI. However, other than blood-brain barrier (BBB) disruption, the influence of oxidative stress on microvessels remains to be elucidated. The aim of this study was to investigate the role of oxidative stress on microcirculatory integrity in EBI. SAH was induced in male Sprague-Dawley rats using an endovascular perforation technique. A free radical scavenger, edaravone, was administered prophylactically by intraperitoneal injection. SAH grade, neurological score, brain water content, and BBB permeability were measured at 24 h after SAH induction. In addition, cortical samples taken at 24 h after SAH were analyzed to explore oxidative stress, microvascular mural cell apoptosis, microspasm, and microthrombosis. Edaravone treatment significantly ameliorated neurological deficits, brain edema, and BBB disruption. In addition, oxidative stress-induced modifications and subsequent apoptosis of microvascular endothelial cells and pericytes increased after SAH induction, while the administration of edaravone suppressed this. Consistent with apoptotic cell inhibition, microthromboses were also inhibited by edaravone administration. Oxidative stress plays a pivotal role in the induction of multiple pathological changes in microvessels in EBI. Antioxidants are potential candidates for the treatment of microvascular disturbances after SAH.

Importance and accuracy of intraoperative frozen section diagnosis of the resection margin for effective carmustine wafer implantation.

For effective implantation of carmustine (BCNU) wafers, it is important to determine the order of priority with reference to the intraoperative frozen section diagnosis of the resection margin (IOFM). The accuracy of IOFM and patterns of tumor recurrence with implantation of BCNU wafers were studied retrospectively. Forty-six cases of newly diagnosed malignant glioma were evaluated. Tumors were resected after intraoperative frozen section diagnosis (IOFD). IOFM was performed for resection walls and evaluated on a three-level scale (-, no tumor invasion; 1+, minor cell invasion; 2+, evident cell invasion). The results were used for effective BCNU wafer implantation. The IOFM sections were then thawed, frozen-paraffin marginal (FPM) sections were prepared, and IOFM was evaluated with FPM sections. The accuracy of IOFD grading was compared to that of the formalin fixed paraffin-embedded section and was 76.1%. The accuracy of IOFM was compared with the FPM section in 148 specimens from 42 patients. The IOFM accuracy was 80.4%. BCNU wafers were implanted in 25 patients and there was recurrence in 15. Local recurrence was seen in 40% (6 patients). However, there was no recurrence immediately below the BCNU wafers. With properly performed IOFM, BCNU wafers can be efficiently implanted, and local recurrence immediately below the BCNU wafers can be inhibited.

Intra-arterial oxidative stress correlates negatively with cognitive function and positively with postoperative ischemic lesions in carotid artery stenosis stenting.

BACKGROUND AND PURPOSE: Carotid plaque contains biologically active substances released into the blood during carotid artery stenting (CAS). The main purpose of this prospective study was to analyse sequential changes in oxidative stress during CAS and their relationship to clinical factors. METHODS: Twenty-two consecutive CAS procedures were performed between May 2014 and April 2016. Arterial blood was collected four times: (1) after the sheath insertion without edaravone; (2) pre-angioplasty with edaravone from the carotid artery; (3) after post-stenting angioplasty from an occluded carotid artery; and (4) before sheath removal. Derivatives of reactive oxygen metabolites (d-ROMs) and biological antioxidant potential (BAP) were measured photometrically. The relationship between d-ROMs or BAP and preoperatively investigated biochemical parameters, cognitive function, and number of diffusion-weighted image (DWI) high spot lesions was analysed using one-way ANOVA and the Tukey-Kramer HSD test. RESULTS: The d-ROM values for CAS were 355±58.8 Carratelli Units at sheath insertion, 315±57.2 after edaravone infusion, 328±56.8 after post-stenting angioplasty, and 315±53.0 just before sheath removal. The d-ROM values were reduced significantly after edaravone infusion (P<0.05). The BAP at sheath insertion was reduced significantly according to age (P<0.05). The d-ROMs at sheath insertion correlated negatively with the dementia scale and positively with the post-CAS DWI high spots (1.00±1.07; P<0.05). Other biochemical parameters did not correlate with the d-ROM values or BAP. CONCLUSION: Oxidative stress is correlated negatively with cognitive function and positively with postoperative ischemic lesions. Antioxidant potential decreases with ageing.

Serum CTX levels and histomorphometric analysis in Src versus RANKL knockout mice.

Src knockout (KO) and RANKL KO mice both exhibit near complete osteopetrosis in terms of 3D-bone volume (BV) fraction by micro-CT, whereas the serum CTX concentration of Src KO is apparently normal and that of RANKL KO is 30% of wild-type (WT) despite the fact that they lack osteoclasts. By histomorphometry we found that, whereas eroded surface (ES) and osteoid surface (OS) are zero values in RANKL KO, they are indistinguishable from WT in Src KO; because of marked increase in bone surface (BS), ES/BS and OS/BS of Src KO are 30-40% of WT. While RANKL KO lack both osteoclasts and osteoblasts, Src KO reveal increased numbers of osteoclasts and indistinguishable numbers of osteoblasts compared with WT; again, on the basis of BS, N.Oc/BS is comparable to WT and N.Ob/BS is markedly decreased in Src KO. The apparently increased number of total osteoclasts may be due to increased expression of RANKL found in Src KO bone in vivo. Src has a gene dosage-dependent effect on osteoclast function in vitro, with Src-/- osteoclasts completely lacking bone-resorbing function as determined by CTX release on dentin. Thus, Src KO osteoclasts retain some bone-resorbing function in vivo. The number of osteocytes is proportionally increased in RANKL KO, while Src KO mice have relative osteocyte deficiency, raising the possibility that RANKL and Src has an unrecognized role in osteocyte survival.

Regression of atherosclerosis with apple procyanidins by activating the ATP-binding cassette subfamily A member 1 in a rabbit model.

BACKGROUND AND AIMS: Apple polyphenol contains abundant procyanidins, which have been associated with an anti-atherosclerosis and cholesterol-lowering effect. The aim of this study was to investigate whether apple procyanidins (APCs) feature therapeutic efficacy in terms of regressing atherosclerosis and whether this efficacy is due to mechanisms other than a cholesterol-lowering effect. METHODS: After eight weeks on an atherogenic diet, rabbits were given a normal diet for another eight weeks to normalize the increased serum lipids level. The rabbits in the baseline group were sacrificed at this stage. The control group was subsequently fed a normal diet for eight weeks, while the APCs group was administrated 50 mg/kg/day of APCs in addition to the normal diet. Serum lipids and aortic intimal-medial thickness (IMT) were serially examined, and the resected aorta was examined histologically and through molecular biology. RESULTS: Aortic IMT on ultrasonography and the lipid accumulation area examined using Sudan IV staining were significantly reduced in the APCs group as compared to the control group. Serum lipid profiles were not different between the groups. Immunohistochemistry showed significantly decreased staining of an oxidative stress marker and significantly increased staining of ATP-binding cassette subfamily A member 1 (ABCA1) in the APCs group. Western blotting and RT-PCR also showed increased expression of ABCA1 mRNA and its protein in the APCs group. CONCLUSIONS: This study revealed that APCs administration causes a regression of atherosclerosis. APCs might hold promise as an anti-atherosclerotic agent.

Age-related marrow adipogenesis is linked to increased expression of RANKL.

With advancing age bone marrow is progressively replaced with adipose tissue, accompanied by a concomitant decline in bone mass and strength. The mechanism underlying the increase in marrow fat and bone destruction remains elusive. We found that on the way of adipogenic differentiation of marrow stromal cells, receptor activator for NF-κB ligand (Rankl) expression was induced, concomitantly with a down-regulation of osteoprotegerin, which prompted us to hypothesize that cells at a preadipocyte stage express RANKL. This concept was supported by the findings that the early adipogenic transcription factors C/EBPß and C/EBPδ, but not the late factor peroxisome proliferator-activated receptor γ, bind to the Rankl promoter and stimulate Rankl gene transcription. In fact, when cells isolated from the bone marrow of aging mice were analyzed by flow cytometry, we found that cells expressing the pre-adipocyte marker Pref-1 were RANKL-positive, and the number of these cells was increased with aging, with concomitant down-regulation of osteoprotegerin, and most importantly, that these RANKL(+)/Pref-1(+) marrow cells were capable of generating osteoclasts from bone marrow macrophages. Thus, the capacity of cells at a pre-adipocyte stage to express RANKL via C/EBPß and C/EBPδ and to support osteoclastogenesis may account partly for the co-progression of fatty marrow and bone destruction with aging.

Mineralocorticoid receptor function in bone metabolism and its role in glucocorticoid-induced osteopenia.

Although the mineralocorticoid receptor (MR) is expressed in osteoblasts and osteocytes and frequently co-localizes with the glucocorticoid receptors (GR), its pathophysiological functions in bone remain elusive. We report here that pharmacologic inhibition of MR function with eplerenone resulted in increased bone mass, with stimulation of bone formation and suppression of resorption, while specific genetic deletion of MR in osteoblast lineage cells had no effect. Further, treatment with eplerenone as well as specific deletion of MR in osteocytes ameliorated the cortical bone thinning caused by slow-release prednisolone pellets. Thus, MR may be involved in the deleterious effects of glucocorticoid excess on cortical bone.

Lanthanum carbonate stimulates bone formation in a rat model of renal insufficiency with low bone turnover.

Control of phosphate is important in the management of chronic kidney disease with mineral and bone disorder (CKD-MBD), for which lanthanum carbonate, a non-calcium phosphate-binding agent, has recently been introduced; however, it remains to be determined whether it has any beneficial or deleterious effect on bone remodeling. In the present study, the effects of lanthanum carbonate were examined in an animal model that mimics low turnover bone disease in CKD, i.e., thyroparathyroidectomized (TPTX) and 5/6 nephrectomized (NX) rats undergoing a constant infusion of parathyroid hormone (PTH) and thyroxine injections (TPTX-PTH-5/6NX). Bone histomorphometry at the second lumbar vertebra and tibial metaphysis revealed that both bone formation and resorption were markedly suppressed in the TPTX-PTH-5/6NX model compared with the sham-operated control group, and treatment with lanthanum carbonate was associated with the stimulation of bone formation but not an acceleration of bone resorption. Lanthanum treatment caused a robust stimulation of bone formation with an activation of osteoblasts on the endosteal surface of femoral diaphysis, leading to an increase in cortical bone volume. Thus, lanthanum carbonate has the potential to stimulate bone formation in cases of CKD-MBD with suppressed bone turnover.

Physiological functions of osteoblast lineage and T cell-derived RANKL in bone homeostasis.

The cytokine RANKL is essential for osteoclast development in bone. The cellular sources of RANKL for support of osteoclast generation under various pathophysiological conditions have remained unclear, however. Here we show that inactivation of Rankl specifically in osteoblast lineage cells of mice with the use of an Osterix-Cre transgene results in typical osteopetrosis in the trabecular compartment of the tibia, with the phenotype being progressively less marked in the femur and vertebrae. In contrast to its effects on trabecular bone, RANKL deficiency in osteoblast lineage resulted in thinning of the femoral cortex in association with suppression of bone formation during the modeling process. Ablation of RANKL specifically in T cells resulted in a moderate but significant increase in tibial trabecular bone. Mice with RANKL deficiency in osteoblast lineage were protected from bone loss induced by ovariectomy as well as from joint destruction associated with arthritis, whereas loss of RANKL in T cells did not confer such protection. Finally, inducible deletion of Rankl selectively in the osteoblasts from 6 to 12 weeks of age resulted in an increase in bone mass in association with reduced bone resorption and formation. Our results thus suggest that RANKL produced by osteoblasts contributes to osteoclast development in vivo.

Osteoclast-secreted CTHRC1 in the coupling of bone resorption to formation.

Bone remodeling is characterized by the sequential, local tethering of osteoclasts and osteoblasts and is key to the maintenance of bone integrity. While bone matrix-mobilized growth factors, such as TGF-ß, are proposed to regulate remodeling, no in vivo evidence exists that an osteoclast-produced molecule serves as a coupling factor for bone resorption to formation. We found that CTHRC1, a protein secreted by mature bone-resorbing osteoclasts, targets stromal cells to stimulate osteogenesis. Cthrc1 expression was robustly induced when mature osteoclasts were placed on dentin or hydroxyapatite, and also by increasing extracellular calcium. Cthrc1 expression in bone increased in a high-turnover state (such as that induced by RANKL injections in vivo), but decreased in conditions associated with suppressed bone turnover (such as with aging and after alendronate treatment). Targeted deletion of Cthrc1 in mice eliminated Cthrc1 expression in bone, whereas its deficiency in osteoblasts did not exert any significant effect. Osteoclast-specific deletion of Cthrc1 resulted in osteopenia due to reduced bone formation and impaired the coupling process after resorption induced by RANKL injections, impairing bone mass recovery. These data demonstrate that CTHRC1 is an osteoclast-secreted coupling factor that regulates bone remodeling.

Physiological function of the angiotensin AT1a receptor in bone remodeling.

In order to determine whether the renin-angiotensin system (RAS) has any physiologic function in bone metabolism, mice lacking the gene encoding the major angiotensin II receptor isoform, AT1a, were studied using micro CT scanning, histomorphometric, and biochemical techniques. Three-dimensional (3D) micro CT analysis of the tibial metaphysis revealed that both male and female AT1a knockout mice exhibited an increased trabecular bone volume along with increased trabecular number and connectivity. Histomorphometric analysis of the tibial metaphysis indicated that the parameters of bone formation as well as resorption were increased, which was also supported by elevated serum osteocalcin and carboxy-terminal collagen crosslink (CTX) concentrations in the AT1a-deficient mice. Osteoclastogenesis and osteoblastogenesis assays in ex vivo cultures, however, did not reveal any intrinsic alterations in the differentiation potential of AT1a-deficient cells. Quantitative RT-PCR using RNA isolated from the tibia and femur revealed that the receptor activator of NF-κB ligand (RANKL)/osteoprotegerin (OPG) ratio and the expression of stromal cell-derived factor (SDF)1α were increased, whereas that of SOST was decreased in AT1a-deficient bone, which may account for the increased bone resorption and formation, respectively. AT1a-deficient mice also displayed a lean phenotype with reduced serum leptin levels. They maintained high bone mass with advancing age, and were protected from bone loss induced by ovariectomy. Collectively, the data suggest that RAS has a physiologic function in bone remodeling, and that signaling through AT1a negatively regulates bone turnover and bone mass.

Coordination of PGC-1beta and iron uptake in mitochondrial biogenesis and osteoclast activation.

Osteoclasts are acid-secreting polykaryons that have high energy demands and contain abundant mitochondria. How mitochondrial biogenesis is integrated with osteoclast differentiation is unknown. We found that the transcription of Ppargc1b, which encodes peroxisome proliferator-activated receptor-gamma coactivator 1beta (PGC-1beta), was induced during osteoclast differentiation by cAMP response element-binding protein (CREB) as a result of reactive oxygen species. Knockdown of Ppargc1b in vitro inhibited osteoclast differentiation and mitochondria biogenesis, whereas deletion of the Ppargc1b gene in mice resulted in increased bone mass due to impaired osteoclast function. We also observed defects in PGC-1beta-deficient osteoblasts. Owing to the heightened iron demand in osteoclast development, transferrin receptor 1 (TfR1) expression was induced post-transcriptionally via iron regulatory protein 2. TfR1-mediated iron uptake promoted osteoclast differentiation and bone-resorbing activity, associated with the induction of mitochondrial respiration, production of reactive oxygen species and accelerated Ppargc1b transcription. Iron chelation inhibited osteoclastic bone resorption and protected against bone loss following estrogen deficiency resulting from ovariectomy. These data establish mitochondrial biogenesis orchestrated by PGC-1beta, coupled with iron uptake through TfR1 and iron supply to mitochondrial respiratory proteins, as a fundamental pathway linked to osteoclast activation and bone metabolism.

Activation of renin-angiotensin system induces osteoporosis independently of hypertension.

Hypertension and osteoporosis are two major age-related disorders; however, the underlying molecular mechanism for this comorbidity is not known. The renin-angiotensin system (RAS) plays a central role in the control of blood pressure and has been an important target of antihypertensive drugs. Using a chimeric RAS model of transgenic THM (Tsukuba hypertensive mouse) expressing both the human renin and human angiotensinogen genes, we showed in this study that activation of RAS induces high turnover osteoporosis with accelerated bone resorption. Transgenic mice that express only the human renin gene were normotensive and yet exhibited a low bone mass, suggesting that osteoporosis occurs independently of the development of hypertension per se. Ex vivo cultures showed that angiotensin II (AngII) acted on osteoblasts and not directly on osteoclast precursor cells and increased osteoclastogenesis-supporting cytokines, RANKL and vascular endothelial growth factor (VEGF), thereby stimulating the formation of osteoclasts. Knockdown of AT2 receptor inhibited the AngII activity, whereas silencing of the AT1 receptor paradoxically enhanced it, suggesting a functional interaction between the two AngII receptors on the osteoblastic cell surface. Finally, treatment of THM mice with an ACE inhibitor, enalapril, improved osteoporosis and hypertension, whereas treatment with losartan, an angiotensin receptor blockers specific for AT1, resulted in exacerbation of the low bone mass phenotype. Thus, blocking the synthesis of AngII may be an effective treatment of osteoporosis and hypertension, especially for those afflicted with both conditions.

Orphan nuclear receptor Nur77 accelerates the initial phase of adipocyte differentiation in 3T3-L1 cells by promoting mitotic clonal expansion.

Nur77 is an orphan member of the nuclear receptor superfamily that is expressed in various types of cells and mediates diverse biological processes. Although Nur77 mRNA is induced in the early stage of adipogenesis of 3T3-L1 cells, its roles are not known. To address this issue, we closely inspected the expression of Nur77 mRNA and protein during differentiation of 3T3-L1 cells. Nur77 was induced rapidly and transiently at both mRNA and protein levels only in the initial phase of differentiation induction, and localized almost exclusively in the nuclei. Isobutylmethylxanthine was essential for the induction of Nur77 protein, acting by at least in part protecting the protein from rapid degradation by proteasome. Nur77 siRNA resulted in delayed adipogenesis in 3T3-L1, accompanied by retarded mitotic clonal expansion. These effects were mediated at least partly by decreased expression of cyclins D and E. Constitutive expression of Nur77 inhibited adipogenesis of 3T3-L1, accompanied by enhanced expression of cyclin D1 and prolonged mitotic clonal expansion. Moreover, constitutive expression of Nur77 inhibited, but transient induction of Nur77 promoted, adipogenesis in NIH-3T3 cells. These results suggest that Nur77 accelerates adipocyte differentiation by regulating cell cycle progression and the rapid and transient induction is crucial for its action.