Interaction between TRPV1-expressing neurons in the hypothalamus.

Transient receptor potential vanilloid type 1 (TRPV1) is a ligand-gated ion channel expressed in the peripheral and central nervous systems. TRPV1-dependent mechanisms take part in a wide range of physiological and pathophysiological pathways including the regulation of homeostatic functions. TRPV1 expression in the hypothalamus has been described as well as evidence that TRPV1-dependent excitatory inputs to hypothalamic preautonomic neurons are diminished in diabetic conditions. Here we aimed to determine the functional expression of TRPV1 in two hypothalamic nuclei known to be involved in the central control of metabolism and to test the hypothesis that TRPV1-expressing neurons receive TRPV1-expressing inputs. A mouse model (TRPV1Cre/tdTom) was generated to identify TRPV1-expressing cells and determine the cellular properties of TRPV1-expressing neurons in adult mice. Our study demonstrated the functional expression of TRPV1 in the dorsomedial hypothalamic nucleus and paraventricular nucleus in adult mice. Our findings revealed that a subset of TRPV1Cre/tdTom neurons receive TRPV1-expressing excitatory inputs, indicating direct interaction between TRPV1-expressing neurons. In addition, astrocytes likely play a role in the modulation of TRPV1-expressing neurons. In summary, this study identified specific hypothalamic regions where TRPV1 is expressed and functional in adult mice and the existence of direct connections between TRPV1Cre/tdTom neurons. NEW & NOTEWORTHY Transient receptor potential vanilloid type 1 (TRPV1) is expressed in the hypothalamus, and TRPV1-dependent regulation of preautonomic neurons is decreased in hyperglycemic conditions. Our study demonstrated functional expression of TRPV1 in two hypothalamic nuclei involved in the control of energy homeostasis. Our results also revealed that a subset of TRPV1-expressing neurons receive TRPV1-expressing excitatory inputs. These findings suggest direct interaction between TRPV1-expressing neurons.

Canagliflozin Prevents Intrarenal Angiotensinogen Augmentation and Mitigates Kidney Injury and Hypertension in Mouse Model of Type 2 Diabetes Mellitus.

BACKGROUND: Hypertension and renal injury are common complications of type 2 diabetes mellitus (T2DM). Hyperglycemia stimulates renal proximal tubular angiotensinogen (AGT) expression via elevated oxidative stress contributing to the development of high blood pressure and diabetic nephropathy. The sodium glucose cotransporter 2 (SGLT2) in proximal tubules is responsible for the majority of glucose reabsorption by renal tubules. We tested the hypothesis that SGLT2 inhibition with canagliflozin (CANA) prevents intrarenal AGT augmentation and ameliorates kidney injury and hypertension in T2DM. METHODS: We induced T2DM in New Zealand obese mice with a high fat diet (DM, 30% fat) with control mice receiving regular fat diet (ND, 4% fat). When DM mice exhibited > 350 mg/dL blood glucose levels, both DM- and ND-fed mice were treated with 10 mg/kg/day CANA or vehicle by oral gavage for 6 weeks. We evaluated intrarenal AGT, blood pressure, and the development of kidney injury. RESULTS: Systolic blood pressure in DM mice (133.9 ± 2.0 mm Hg) was normalized by CANA (113.9 ± 4.0 mm Hg). CANA treatment ameliorated hyperglycemia-associated augmentation of renal AGT mRNA (148 ± 21 copies/ng RNA in DM, and 90 ± 16 copies/ng RNA in DM + CANA) and protein levels as well as elevation of urinary 8-isoprostane levels. Tubular fibrosis in DM mice (3.4 ± 0.9-fold, fibrotic score, ratio to ND) was suppressed by CANA (0.9 ± 0.3-fold). Furthermore, CANA attenuated DM associated increased macrophage infiltration and cell proliferation in kidneys of DM mice. CONCLUSIONS: CANA prevents intrarenal AGT upregulation and oxidative stress and which may mitigate high blood pressure, renal tubular fibrosis, and renal inflammation in T2DM.

Overactivity of Liver-Related Neurons in the Paraventricular Nucleus of the Hypothalamus: Electrophysiological Findings in db/db Mice.

Preautonomic neurons in the paraventricular nucleus (PVN) of the hypothalamus play a large role in the regulation of hepatic functions via the autonomic nervous system. Activation of hepatic sympathetic nerves increases glucose and lipid metabolism and contributes to the elevated hepatic glucose production observed in the type 2 diabetic condition. This augmented sympathetic output could originate from altered activity of liver-related PVN neurons. Remarkably, despite the importance of the brain-liver pathway, the cellular properties of liver-related neurons are not known. In this study, we provide the first evidence of overall activity of liver-related PVN neurons. Liver-related PVN neurons were identified with a retrograde, trans-synaptic, viral tracer in male lean and db/db mice and whole-cell patch-clamp recordings were conducted. In db/db mice, the majority of liver-related PVN neurons fired spontaneously; whereas, in lean mice the majority of liver-related PVN neurons were silent, indicating that liver-related PVN neurons are more active in db/db mice. Persistent, tonic inhibition was identified in liver-related PVN neurons; although, the magnitude of tonic inhibitory control was not different between lean and db/db mice. In addition, our study revealed that the transient receptor potential vanilloid type 1-dependent increase of excitatory neurotransmission was reduced in liver-related PVN neurons of db/db mice. These findings demonstrate plasticity of liver-related PVN neurons and a shift toward excitation in a diabetic mouse model. Our study suggests altered autonomic circuits at the level of the PVN, which can contribute to autonomic dysfunction and dysregulation of neural control of hepatic functions including glucose metabolism.SIGNIFICANCE STATEMENT A growing body of evidence suggests the importance of the autonomic control in the regulation of hepatic metabolism, which plays a major role in the development and progression of type 2 diabetes mellitus. Despite the importance of the brain-liver pathway, the overall activity of liver-related neurons in control and diabetic conditions is not known. This is a significant gap in knowledge, which prevents developing strategies to improve glucose homeostasis via altering the brain-liver pathway. One of the key findings of our study is the overall shift toward excitation in liver-related hypothalamic neurons in the diabetic condition. This overactivity may be one of the underlying mechanisms of elevated sympathetic activity known in metabolically compromised patients and animal models.

The effects of an educational program for non-physician health care providers regarding fertility preservation.

PURPOSE: The American Society of Clinical Oncology (ASCO) Clinical Practice guidelines recommend that physicians, nurses, social workers, and other health care providers should be prepared to discuss the risk of infertility with patients. We conducted an educational program for non-physician health care providers regarding fertility preservation and evaluated the effects of the educational program. METHODS: The 4-h educational program consisted of lectures about infertility as a potential risk of cancer treatment, fertility preservation, and psychosocial support. Knowledge, confidence, institutional change, and self-practice were assessed pre-program, immediately post-program, and 6 months post-program. RESULTS: Of 124 participants who joined the program, 74 completed and returned the follow-up survey 6 months after the program. Sixty-one percent of the participants were nurses, 27% were social workers, and 4% were psychologists. The scores for confidence and knowledge increased between pre- and immediate post-program periods (p < 0.01), and between pre- and 6-month post-program periods (p < 0.01). The knowledge score was 52, 76, and 71% at the 3 points respectively. The participants became more likely to disseminate fertility preservation counseling at their institutions (p < 0.01) and use informational resources (p < 0.01). Overall, self-practice and institutional support did not change. CONCLUSIONS: The study revealed that this educational program is applicable for non-physicians to learn about fertility preservation. The participants improved significantly in confidence and knowledge, but not in counseling skills.

Macrophage-derived IL-6 contributes to ANG II-mediated angiotensinogen stimulation in renal proximal tubular cells.

The development of ANG II-dependent hypertension involves increased infiltration of macrophages (MΦ) and T cells into the kidney and the consequent elevation of intrarenal cytokines including IL-6, which facilitates the progression of hypertension and associated kidney injury. Intrarenal renin-angiotensin system (RAS) activation, including proximal tubular angiotensinogen (AGT) stimulation, has also been regarded as a cardinal mechanism contributing to these diseases. However, the interaction between immune cells and intrarenal RAS activation has not been fully delineated. Therefore, the present study investigated whether ANG II-treated MΦ induce AGT upregulation in renal proximal tubular cells (PTCs). MΦ were treated with 0-10(-6) M ANG II for up to 48 h. PTCs were incubated with the collected medium from MΦ. In ANG II-treated MΦ, IL-6 mRNA and protein levels were increased (1.86 ± 0.14, protein level, ratio to control); moreover, IL-6 levels were higher than TNF-α and IL-1ß in culture medium isolated from ANG II-treated MΦ. Elevated AGT expression (1.69 ± 0.04, ratio to control) accompanied by phosphorylated STAT3 were observed in PTCs that received culture medium from ANG II-treated MΦ. The addition of a neutralizing IL-6 antibody to the collected medium attenuated phosphorylation of STAT3 and AGT augmentation in PTCs. Furthermore, a JAK2 inhibitor also suppressed STAT3 phosphorylation and AGT augmentation in PTCs. These results demonstrate that ANG II-induced IL-6 elevation in MΦ enhances activation of the JAK-STAT pathway and consequent AGT upregulation in PTCs, suggesting involvement of an immune response in driving intrarenal RAS activity.

Quantification of intact plasma AGT consisting of oxidized and reduced conformations using a modified ELISA.

The pleiotropic actions of the renin-angiotensin system (RAS) depend on the availability of angiotensinogen (AGT) which generates angiotensin I (ANG I) when cleaved by renin. Thus, quantification of the intact AGT (iAGT) concentrations is important to evaluate the actual renin substrate available. The iAGT conformation exists as oxidized AGT (oxi-AGT) and reduced AGT (red-AGT) in a disulfide bond, and oxi-AGT has a higher affinity for renin, which may exacerbate RAS-associated diseases. Accordingly, we determined iAGT, oxi-AGT, and red-AGT levels in plasma from rats and mice. Blood samples were obtained by cardiac puncture and then immediately mixed with an inhibitor solution containing a renin inhibitor. Total AGT (tAGT) levels were measured by tAGT ELISA which detects both cleaved and iAGT. iAGT levels were determined by iAGT ELISA which was found to only detect red-AGT. Thus, it was necessary to treat samples with dithiothreitol, a reducing agent, to quantify total iAGT concentration. tAGT levels in rat and mouse plasma were 1,839 ± 139 and 1,082 ± 77 ng/ml, respectively. iAGT levels were 53% of tAGT in rat plasma but only 22% in mouse plasma, probably reflecting the greater plasma renin activity in mice. The ratios of oxi-AGT and red-AGT were ∼4:1 (rat) and 16:1 (mouse). Plasma iAGT consists of oxi-AGT and red-AGT, suggesting that oxidative stress can influence ANG I generation by the AGT conformation switch. Furthermore, the lower availability of plasma iAGT in mice suggests that it may serve as a limiting factor in ANG I formation in this species.

Increased angiotensinogen expression, urinary angiotensinogen excretion, and tissue injury in nonclipped kidneys of two-kidney, one-clip hypertensive rats.

In angiotensin II (ANG II)-dependent hypertension, there is an angiotensin type 1 receptor-dependent amplification mechanism enhancing intrarenal angiotensinogen (AGT) formation and secretion in the tubular fluid. To evaluate the role of increased arterial pressure, AGT mRNA, protein expression, and urinary AGT (uAGT) excretion and tissue injury were assessed in both kidneys of two-kidney, one-clip Sprague-Dawley hypertensive rats subjected to left renal arterial clipping (0.25-mm gap). By 18-21 days, systolic arterial pressure increased to 180 ± 3 mmHg, and uAGT increased. Water intake, body weights, 24-h urine volumes, and sodium excretion were similar. In separate measurements of renal function in anesthetized rats, renal plasma flow and glomerular filtration rate were similar in clipped and nonclipped kidneys and not different from those in sham rats, indicating that the perfusion pressure to the clipped kidneys remained within the autoregulatory range. The nonclipped kidneys exhibited increased urine flow and sodium excretion. The uAGT excretion was significantly greater in nonclipped kidneys compared with clipped and sham kidneys. AGT mRNA was 2.15-fold greater in the nonclipped kidneys compared with sham (1.0 ± 0.1) or clipped (0.98 ± 0.15) kidneys. AGT protein levels were also greater in the nonclipped kidneys. The nonclipped kidneys exhibited greater glomerular expansion and immune cell infiltration, medullary fibrosis, and cellular proliferation than the clipped kidneys. Because both kidneys have elevated ANG II levels, the greater tissue injury in the nonclipped kidneys indicates that an increased arterial pressure synergizes with increased intrarenal ANG II to stimulate AGT production and exert greater renal injury.

Brain stem as a target site for the metabolic side effects of olanzapine.

Olanzapine, an atypical antipsychotic, is widely prescribed for the treatment of schizophrenia and bipolar disorder despite causing undesirable metabolic side effects. A variety of mechanisms and brain sites have been proposed as contributors to the side effects; however, the role of the dorsal motor nucleus of the vagus nerve (DMV), which plays a crucial role in the regulation of subdiaphragmatic organs and thus governs energy and glucose homeostasis, is largely unknown. Identifying the effect of olanzapine on the excitability of DMV neurons in both sexes is thus crucial to understanding possible underlying mechanisms. Whole cell patch-clamp electrophysiological recordings were conducted in stomach- and liver-related DMV neurons identified with retrograde viral tracers and in random DMV neurons. The effect of olanzapine on the neuronal excitability of DMV neurons both in male and female mice was established. Our data demonstrate that olanzapine hyperpolarizes the DMV neurons in both sexes and this effect is reversible. The hyperpolarization is associated with decreased firing rate and input resistance. Olanzapine also decreases the excitability of a subset of stomach- and liver-related DMV neurons. Our study demonstrates that olanzapine has a powerful effect on DMV neurons in both sexes, indicating its ability to reduce vagal output to the subdiaphragmatic organs, which likely contributes to the metabolic side effects observed in both humans and experimental models. These findings suggest that the metabolic side effects of olanzapine may partially originate in the DMV.

ROCK/NF-κB axis-dependent augmentation of angiotensinogen by angiotensin II in primary-cultured preglomerular vascular smooth muscle cells.

In angiotensin II (ANG II)-dependent hypertension, the augmented intrarenal ANG II constricts the renal microvasculature and stimulates Rho kinase (ROCK), which modulates vascular contractile responses. Rho may also stimulate angiotensinogen (AGT) expression in preglomerular vascular smooth muscle cells (VSMCs), but this has not been established. Therefore, the aims of this study were to determine the direct interactions between Rho and ANG II in regulating AGT and other renin-angiotensin system (RAS) components and to elucidate the roles of the ROCK/NF-κB axis in the ANG II-induced AGT augmentation in primary cultures of preglomerular VSMCs. We first demonstrated that these preglomerular VSMCs express renin, AGT, angiotensin-converting enzyme, and ANG II type 1 (AT1) receptors. Furthermore, incubation with ANG II (100 pmol/l for 24 h) increased AGT mRNA (1.42 ± 0.03, ratio to control) and protein (1.68 ± 0.05, ratio to control) expression levels, intracellular ANG II levels, and NF-κB activity. In contrast, the ANG II treatment did not alter AT1a and AT1b mRNA levels in the cells. Treatment with H-1152 (ROCK inhibitor, 10 nmol/l) and ROCK1 small interfering (si) RNA suppressed the ANG II-induced AGT augmentation and the upregulation and translocalization of p65 into nuclei. Functional studies showed that ROCK exerted a greater influence on afferent arteriole responses to ANG II in rats subjected to chronic ANG II infusions. These results indicate that ROCK is involved in NF-κB activation and the ROCK/NF-κB axis contributes to ANG II-induced AGT upregulation, leading to intracellular ANG II augmentation.

Interferon-γ biphasically regulates angiotensinogen expression via a JAK-STAT pathway and suppressor of cytokine signaling 1 (SOCS1) in renal proximal tubular cells.

Renal inflammation modulates angiotensinogen (AGT) production in renal proximal tubular cells (RPTCs) via inflammatory cytokines, including interleukin-6, tumor necrosis factor α, and interferon-γ (IFN-γ). Among these, the effects of IFN-γ on AGT regulation in RPTCs are incompletely delineated. This study aimed to elucidate mechanisms by which IFN-γ regulates AGT expression in RPTCs. RPTCs were incubated with or without IFN-γ up to 48 h. AGT expression, STAT1 and STAT3 activities, and SOCS1 expression were evaluated. RNA interference studies against STAT1, SOCS1, and STAT3 were performed to elucidate a signaling cascade. IFN-γ decreased AGT expression at 6 h (0.61±0.05, ratio to control) and 12 h (0.47±0.03). In contrast, longer exposure for 24 and 48 h increased AGT expression (1.76±0.18, EC(50)=3.4 ng/ml, and 1.45±0.08, respectively). IFN-γ treatment for 6 h strongly induced STAT1 phosphorylation and SOCS1 augmentation, and decreased STAT3 activity. However, STAT1 phosphorylation and SOCS1 augmentation waned at 24 h, while STAT3 activity increased. RNA interference studies revealed that activation of STAT1-SOCS1 axis decreased STAT3 activity. Thus, IFN-γ biphasically regulates AGT expression in RPTCs via STAT3 activity modulated by STAT1-SOCS1 axis, suggesting the STAT1-SOCS1 axis is important in IFN-γ-induced activation of the intrarenal renin-angiotensin system.

Safety and Effectiveness of Apixaban Versus Warfarin in Japanese Patients with Nonvalvular Atrial Fibrillation Stratified by Renal Function: A Retrospective Cohort Study.

BACKGROUND: We previously conducted a retrospective cohort study using chart review of oral anticoagulant-naïve Japanese patients with nonvalvular atrial fibrillation (NVAF) that assessed the risk of major bleeding and stroke/systemic embolism (SE) events of apixaban versus warfarin. METHODS: In this subgroup analysis, we compared the risk of major bleeding and stroke/SE events by stratifying patients into four subgroups matched 1:1 using propensity score matching (PSM) according to baseline creatinine clearance (CrCl; mL/min): ≥ 15 to < 30, ≥ 30 to < 50, ≥ 50 to < 80, and ≥ 80. RESULTS: Of the 7074 patients in the apixaban group and 4998 in the warfarin group eligible for inclusion in the analysis, 4385 were included in each group after PSM. Incidence rates of major bleeding and stroke/SE events were generally lower with apixaban versus warfarin across the CrCl subgroups. When all patients with a CrCl change of < 0 mL/min per year during the study period (apixaban, n = 3871; warfarin, n = 2635) were stratified into four subgroups based on the magnitude of CrCl decline (median CrCl change [mL/min] per year: - 1.09, - 3.48, - 7.54, and - 36.92 for apixaban, and - 1.10, - 3.65, - 7.85, and - 40.40 for warfarin), the incidence rates of major bleeding and stroke/SE events generally increased with an increasing CrCl decline per year in both groups. CONCLUSIONS: In Japanese patients with NVAF, the safety and effectiveness of apixaban and warfarin were consistent across different renal subgroups, including those with severe renal impairment. Our results highlight the importance of monitoring renal function variations over time in patients with NVAF. GOV IDENTIFIER: NCT03765242.

[Investigation the contents of employment consultation and support in a cancer center hospital].

OBJECTIVES: This study aimed to analyze and categorize the actual situation of employment consultation and support according to consultation times or employment status at the Consultation Support Center of the National Cancer Center Hospital of Japan. METHODS: We retrospectively analyzed the patient backgrounds, consultation contents, and the number of employment consultation cases conducted at the Consultation Support Center of the National Cancer Center Hospital during a 6-month period from May to December 2018. RESULTS: During the study period, 117 patients (male: female = 46:71) visited the Consultation Support Center. The median age of patients was 48 years old. The most common primary cancer site was the breast in 28 patients followed by the lung in 16 patients, and then gynecologic cancer in 10 patients. The most common cancer treatment was chemotherapy in 53 patients (45.3%), and 12 patients (10.2%) were recurrent patients. Fifty-two patients were in regular employment, 24 were unemployed, 17 were of unknown employment status, 16 were in non-regular employment, and 8 were classified/categorized as other. In terms of working status, 40 were on leave, 35 were working, 15 were seeking work, 8 were unemployed, and 19 were categorized as other. The median number of consultations was 1 (1,11). The content of consultations was the social security system in 44 cases (37.6%) job seeking in 24 cases (20.5%), how to inform the workplace in 14 cases (12%), and workplace environment adjustment in 13 cases (11.1%). CONCLUSIONS: We conducted a survey on the actual status of employment consultation in a cancer center hospital. The majority of consultations were completed in one session. In terms of the content of consultations, there was a high need for consultations on the social security system and job seeking. Further study is needed on the characteristics of employment consultations according to employment status and other attributes.

Tumor necrosis factor-{alpha} suppresses angiotensinogen expression through formation of a p50/p50 homodimer in human renal proximal tubular cells.

Angiotensinogen (AGT) expression in renal proximal tubular cells (RPTCs) and intrarenal tumor necrosis factor-α (TNF-α) levels are increased in hypertension and renal diseases However, the contribution of TNF-α to AGT expression in RPTCs has not been established. Therefore, the objective of the present study was to determine influence of TNF-α on AGT expression in RPTCs. Human kidney-2 (HK-2) cells, immortalized human RPTCs, were treated with several concentrations of TNF-α up to 24 h. AGT mRNA and protein expression were evaluated by RT-PCR and ELISA, respectively. Activation of nuclear factor-κB (NF-κB) by TNF-α was evaluated by Western blot analysis, immunocytochemistry, and electrophoretic mobility shift assay (EMSA). TNF-α suppressed AGT mRNA expression in a dose- and time-dependent manner. Maximum AGT mRNA reduction was caused by 40 ng/ml of TNF-α (0.52 ± 0.09, ratio to control, at 24 h) and at 24 h (0.66 ± 0.05, ratio to control, by 10 ng/ml TNF-α). TNF-α reduced AGT protein accumulation in the medium between 8 and 24 h (0.62 ± 0.13 by 40 ng/ml TNF-α, ratio to control). TNF-α activated and induced translocalization of p50 and p65, which are NF-κB subunits. Elevated formation of p50/p65 and p50/p50 dimers by TNF-α were observed by EMSA and supershift assay. Gene silencing of p50, but not p65, attenuated the effect of TNF-α on reduction of AGT expression in RPTCs. These results indicate that TNF-α suppresses AGT expression through p50/p50 homodimer formation in human RPTCs, suggesting a possible counteracting mechanism that limits excessive intrarenal AGT production.

STAT1 regulates interferon-γ-induced angiotensinogen and MCP-1 expression in a bidirectional manner in primary cultured mesangial cells.

OBJECTIVE: Intrarenal interferon-γ significantly contributes to the development of glomerular injury in which angiotensinogen and monocyte chemoattractant protein 1 levels are elevated. However, the exact nature of the role that interferon-γ plays in regulating angiotensinogen and monocyte chemoattractant protein 1 expression has not been fully delineated. Therefore, the aim of this study was to investigate the role that interferon-γ plays in angiotensinogen and monocyte chemoattractant protein 1 expression. METHODS: Primary cultured rat mesangial cells were treated with 0-20 ng/mL interferon-γ for 2, 8 or 24 hours. Expression levels of angiotensinogen, monocyte chemoattractant protein 1, suppressors of cytokine signaling 1, an intracellular suppressor of Janus kinase-signal transducers and activators of transcription signaling and activity of the Janus kinase-signal transducers and activators of transcription pathway were evaluated by reverse transcriptase polymerase chain reaction and western blot analysis. RESULTS: Interferon-γ increased angiotensinogen expression in mesangial cells with maximal augmentation observed following 5 ng/mL interferon-γ at 8 hours of treatment (1.87 ± 0.05, mRNA, relative ratio). Further increases were reduced or absent using higher concentrations of interferon-γ. Following treatments, monocyte chemoattractant protein 1 expression was induced in a linear dose-dependent manner (6.85 ± 0.62-fold by 20 ng/mL interferon-γ at 24 hours). In addition, interferon-γ induced STAT1 phosphorylation and suppressors of cytokine signaling 1 expression in a linear dose-dependent manner. The suppression of STAT1 and suppressors of cytokine signaling 1 expression by small interference RNAs facilitated an increase in interferon-γ-induced angiotensinogen expression, indicating that these two factors negatively regulate angiotensinogen expression. In contrast, the increase in interferon-γ-induced monocyte chemoattractant protein 1 expression was attenuated in STAT1-deficient mesangial cells, suggesting that STAT1 positively regulates monocyte chemoattractant protein 1 expression in mesangial cells. CONCLUSION: These results demonstrate that while interferon-γ increases both angiotensinogen and monocyte chemoattractant protein 1 expression, STAT1 plays an opposing role in the regulation of each factor in mesangial cells.

Role of activated intrarenal reactive oxygen species and renin-angiotensin system in IgA nephropathy model mice.

1. Using HIGA (high IgA of ddY) mice as an IgA nephropathy model and BALB/c mice as controls, we demonstrated that reactive oxygen species (ROS) and the renin-angiotensin system (RAS) were activated in kidneys of HIGA mice. However, it was difficult to establish an association between renal damage and changes in ROS and the RAS. Therefore, the present study was performed to determine whether renal injury is associated with changes in ROS and the RAS in HIGA mice. 2. Male HIGA mice were divided into four groups of 10 each: (i) untreated mice (HIGA + null); (ii) mice treated with the angiotensin AT(1) receptor antagonist olmesartan (5 mg/kg per day; HIGA + OLM); (iii) mice treated with the superoxide dismutase mimetic tempol (50 mg/kg per day; HIGA + Tempol); and (iv) mice treated with RAS-independent antihypertensive drugs (30 mg/kg per day hydralazine, 0.6 mg/kg per day reserpine and 12 mg/kg per day hydrochlorothiazide; HIGA + HRH). Mice were treated for 5 weeks. 3. Systolic blood pressure decreased significantly in the HIGA + OLM and HIGA + HRH groups, but not in the HIGA + Tempol group, compared with HIGA + null mice. The expression of two ROS markers (4-hydroxy-2-nonenal and heme oxygenase-1) and angiotensin II as a marker of the RAS decreased significantly in HIGA + OLM and HIGA + Tempol mice, but not in HIGA + HRH mice, compared with HIGA + null mice. As a marker of renal damage, mesangial matrix expansion and the desmin-positive area decreased significantly in the HIGA + OLM and HIGA + Tempol groups, but not in HIGA + HRH group, compared with the HIGA + null group. 4. These data suggest that intrarenal ROS and RAS activation play a pivotal role in the development of IgA nephropathy model mice, from the early phase, independent of blood pressure.

Activation of reactive oxygen species and the renin-angiotensin system in IgA nephropathy model mice.

1. Although IgA nephropathy is the most common form of primary glomerulopathy, the detailed mechanisms underlying its development remain uncertain. 2. In the present study, we used male high IgA strain of ddY (HIGA) mice as the IgA nephropathy model and age-matched male BALB/c mice as the control. Recent studies have demonstrated that reactive oxygen species (ROS)-dependent enhancement of the renin-angiotensin system (RAS) plays a potential role in the development and progression of renal injury. Therefore, in the present study we periodically measured the systolic blood pressure (SBP) of mice over the period 21-25 weeks of age and estimated markers for ROS, RAS and renal damage after mice had been killed at 25 weeks of age. 3. Markers for ROS (urinary 8-isoprostane excretion and renal 4-hydroxy-2-nonenal accumulation), RAS (renal angiotensinogen protein expression, urinary angiotensinogen excretion and renal angiotensin II) and renal damage (desmin-positive area and urinary protein excretion), as well as SBP, were significantly increased in HIGA mice compared with control BALB/c mice. 4. The data suggest that both ROS and the RAS are activated at an early phase in IgA nephropathy model mice.

Advanced Glycation End Products Stimulate Angiotensinogen Production in Renal Proximal Tubular Cells.

BACKGROUND: Elevated advanced glycation end products (AGE) in diabetes mellitus (DM) are implicated in the progression of DM-associated tissue injury, including diabetic nephropathy. The intrarenal renin-angiotensin system, in particular augmentation of angiotensinogen (AGT) in proximal tubular cells (PTC), plays a crucial role in the development of diabetic nephropathy. This study investigated hypothesis that AGE stimulates AGT production in PTC. MATERIALS AND METHODS: Urinary AGT and AGE levels in streptozotocin-induced DM mice were measured by enzyme-linked immunosorbent assays. AGT expression and secretion were evaluated in cultured rat PTC receiving 0-200 µg/ml AGE-BSA treatments for 24 hours. Furthermore, intracellular signaling pathways activated by AGE were elucidated. RESULTS: DM mice exhibited greater urinary AGT and AGE levels compared to control mice (AGT: 21.6 ± 5.5 ng/day vs. 190.1 ± 57.8 ng/day, AGE: 139.1 ± 21.6 µg/day vs. 332.8 ± 102.7 µg/day). In cultured PTC, treatment with AGE-BSA enhanced AGT mRNA expression (3.43 ± 0.11-fold compared to control), intracellular AGT protein levels (3.60 ± 0.38-fold), and secreted AGT levels (2.11 ± 0.18-fold). On the other hand, AGT levels were not altered in PTC receiving nonglycated BSA. Recombinant soluble AGE receptor, which competes with endogenous AGE receptor, diminished the AGE-induced AGT upregulation, suggesting that AGE-BSA stimulates AGT expression via activation of the AGE receptor. Enhanced phosphorylation of ERK1/2 and c-Jun, but not p38 MAP kinase, were observed in AGE-BSA-treated PTC. AGE-induced AGT augmentation was attenuated by an ERK inhibitor. CONCLUSIONS: The findings indicate that AGE enhances proximal tubular AGT expression via ERK1/2, which can exacerbate the development of diabetic related kidney injury.

A Content Analysis of Multidimensional Support Needs Regarding Fertility Among Cancer Patients: How Can Nonphysician Health Care Providers Support?

PURPOSE: This study aimed to identify support needs among young cancer patients regarding fertility-related issues to describe multidimensional support provided by nonphysician health care providers. METHODS: Participants were cancer patients and their families who contacted the Hotline for Cancer Treatment and Reproduction at National Cancer Center Hospital in Japan. Medical charts were analyzed through content analysis. RESULTS: A total of 51 participants met the inclusion criteria, of which 32 cases (63%) involved patients themselves, 13 (25%) patients' family members, and 6 (12%) both patients and their families. About patients' demographic status, 40% of the patients were female and 28% were in their thirties. Gynecological and breast cancer patients were the majority, and 24 patients (47%) had not yet started cancer treatment. As a result of content analysis regarding support needs, 9 categories and 55 codes were extracted. The categories included information about risk of infertility, information about means to maintain reproductive function, referral to specialists, support for economic burden, support for worry about cancer progression, support for psychological distress upon facing the risk of infertility, support for communication with oncologists, support for family relationships, and decisional aid. CONCLUSIONS: This study suggests that nonphysician health care providers should acquire knowledge about fertility preservation and provide psychological support within their specialties.

Possible involvement of Rho-kinase in aldosterone-induced vascular smooth muscle cell remodeling.

There is increasing evidence supporting potential roles of aldosterone in the pathogenesis of vascular injury. The present study aimed to determine the involvement of Rho-kinase in aldosterone-induced vascular smooth muscle cell (VSMC) remodeling. In cultured rat VSMC, the effects of aldosterone on Rho-kinase activity, the reorganization of the cytoskeleton and cellular migration were examined. Aldosterone (1 nmol/L) significantly increased phosphorylation of myosin phosphate target subunit-1 (MYPT1), a marker of Rho-kinase activity, and the amount of GTP-Rho with a peak at 90 min in VSMC. Aldosterone also stimulated VSMC stress fiber formation and migration. These effects of aldosterone were markedly attenuated by pretreatment with eplerenone (10 micromol/L), a selective mineralocorticoid receptor antagonist, or Y27632 (10 micromol/L), a specific Rho-kinase inhibitor. These findings indicate that Rho-kinase is involved in the pathogenesis of aldosterone-induced VSMC remodeling.

Sequential activation of the reactive oxygen species/angiotensinogen/renin-angiotensin system axis in renal injury of type 2 diabetic rats.

1. The present study was performed to test the hypothesis that the reactive oxygen species (ROS)-angiotensinogen (AGT)-renin angiotensin system (RAS) axis is sequentially activated in the development of diabetic nephropathy in Zucker diabetic fatty (ZDF) obese rats. 2. Genetic pairs of male ZDF obese and control ZDF lean rats (n = 12 of each species) were killed every 3 weeks from 12 to 21 weeks of age (n = 6 at each time point). 3. The ZDF obese rats developed diabetes mellitus at 12 weeks. At that time, urinary excretion rates of 8-isoprostane were similar between the groups; however, urinary 8-isoprostane levels were significantly increased at 15 weeks in ZDF obese rats compared with controls (36 +/- 6 vs 15 +/- 2 ng/day, respectively). At 15 weeks, protein levels of cortical angiotensinogen were similar between groups; however, cortical angiotensinogen levels were significantly increased at 18 weeks in ZDF obese rats compared with controls (relative ratio of 2.32 +/- 0.21 vs 1.00 +/- 0.20, respectively). At 12 weeks, angiotensin (Ang) II-like immunoreactivity was similar between groups in both the glomeruli and tubules; however, AngII-like immunoreactivity was increased significantly at 21 weeks in ZDF obese rats compared with controls (relative ratios of 1.98 +/- 0.55 vs 1.00 +/- 0.03, respectively, for glomeruli and 1.58 +/- 0.16 vs 1.00 +/- 0.13, respectively, for tubules). Moreover, at 21 weeks, the desmin-positive area in the glomeruli (0.63 +/- 0.08 vs 0.22 +/- 0.05%) and Masson's trichrome stain-positive area in the interstitium (4.97 +/- 0.05 vs 3.18 +/- 0.41%) were significantly increased in ZDF obese rats compared with controls, even though these differences had not been observed earlier. 4. These data suggest that the sequential activation of the ROS-AGT-RAS axis plays an important role in the development of diabetic nephropathy in ZDF obese rats.