Intermittent colonic exoperistalsis for chronic constipation in spinal cord-injured individuals. A long-term structured patient feedback survey to evaluate home care use.

STUDY DESIGN: Structured patient feedback survey evaluating real-world home care use. OBJECTIVES: To assess the long-term effectiveness, tolerability, and satisfaction with the intermittent colonic exoperistalsis (ICE) treatment device MOWOOT in spinal cord-injured (SCI) individuals with chronic constipation. SETTING: Four specialized German hospitals. METHODS: SCI individuals with chronic constipation were invited to use MOWOOT 10-20 min daily and answer a questionnaire about their bowel situation before treatment (feedback 1, F1) and after ≥10 months of use (feedback 2, F2). Collected variables were device use, bowel function effectiveness, chronic constipation symptoms, concomitant use of laxatives and evacuation aids, and satisfaction with bowel function and management, which were compared between time points. At F2, participants reported efficacy, tolerability/side effects, and ease of use. RESULTS: Eleven participants used the device for a mean (SD) of 13.27 (4.03) months. From F1 to F2, mean time per evacuation decreased by 24.5 min (p = 0.0076) and the number of failed attempts to evacuate/week, by 1.05 (p = 0.0354) with a tendency toward increased bowel movements and softer stool consistency, and decreased incomplete bowel movements. Participants experienced decreased difficulty/strain (p = 0.0055), abdominal pain (p = 0.0230), bloating (p = 0.0010), abdominal cramps (p = 0.0019), and spasms (p = 0.0198), without significant changes in the use of laxatives and evacuation aids. Satisfaction with bowel function and management improved (p = 0.0095) and more participants reported being very satisfied/satisfied (p = 0.0300). Most reported tolerability, efficacy, and ease of use as very good/good. CONCLUSION: Long-term in-home ICE treatment improved bowel function and chronic constipation symptoms in SCI individuals, providing clinical benefits to this population. SPONSORSHIP (MOWOOT DEVICES LENDING): 4 M Medical GmbH, Norderstedt, Germany.

The Berlin Bimanual Test for Tetraplegia (BeBiTT): development, psychometric properties, and sensitivity to change in assistive hand exoskeleton application.

BACKGROUND: Assistive hand exoskeletons are promising tools to restore hand function after cervical spinal cord injury (SCI) but assessing their specific impact on bimanual hand and arm function is limited due to lack of reliable and valid clinical tests. Here, we introduce the Berlin Bimanual Test for Tetraplegia (BeBiTT) and demonstrate its psychometric properties and sensitivity to assistive hand exoskeleton-related improvements in bimanual task performance. METHODS: Fourteen study participants with subacute cervical SCI performed the BeBiTT unassisted (baseline). Thereafter, participants repeated the BeBiTT while wearing a brain/neural hand exoskeleton (B/NHE) (intervention). Online control of the B/NHE was established via a hybrid sensorimotor rhythm-based brain-computer interface (BCI) translating electroencephalographic (EEG) and electrooculographic (EOG) signals into open/close commands. For reliability assessment, BeBiTT scores were obtained by four independent observers. Besides internal consistency analysis, construct validity was assessed by correlating baseline BeBiTT scores with the Spinal Cord Independence Measure III (SCIM III) and Quadriplegia Index of Function (QIF). Sensitivity to differences in bimanual task performance was assessed with a bootstrapped paired t-test. RESULTS: The BeBiTT showed excellent interrater reliability (intraclass correlation coefficients > 0.9) and internal consistency (α = 0.91). Validity of the BeBiTT was evidenced by strong correlations between BeBiTT scores and SCIM III as well as QIF. Wearing a B/NHE (intervention) improved the BeBiTT score significantly (p < 0.05) with high effect size (d = 1.063), documenting high sensitivity to intervention-related differences in bimanual task performance. CONCLUSION: The BeBiTT is a reliable and valid test for evaluating bimanual task performance in persons with tetraplegia, suitable to assess the impact of assistive hand exoskeletons on bimanual function.

Quality of life in subjects with upper- and lower-limb spasticity treated with incobotulinumtoxinA.

BACKGROUND: We evaluated quality of life among subjects with upper- and lower-limb spasticity who received escalating doses of incobotulinumtoxinA (total body doses up to 800 U) in the prospective, single-arm, dose-titration TOWER study. METHODS: In this exploratory trial, subjects (N = 155; 18-80 years of age) with upper- and lower-limb spasticity due to cerebral causes who were deemed to require total body doses of up to 800 U incobotulinumtoxinA received three consecutive injection cycles of incobotulinumtoxinA (400, 600, and up to 800 U), each with 12 to 16 weeks' follow-up. QoL was assessed using the EuroQol 5-dimensions questionnaire, three-level (EQ-5D), before and 4 weeks post-injection in each injection cycle and at the end of injection cycle 3. RESULTS: The mean EQ-5D visual analog scale scores of 155 participants continuously improved from study baseline to 4 weeks post-injection in all injection cycles (mean [standard deviation] change 6.7 [14.1], 9.6 [16.3], and 8.6 [17.0] for injection cycles 1, 2, and 3, respectively; p < 0.0001 for all, paired sample t-test). In general, among those with a change in the EQ-5D rating of their condition, the proportion of subjects with 'improvement' was greater than that with 'worsening' for individual EQ-5D dimensions across all injection cycles. At the end of injection cycle 3, the proportion of subjects rating their condition as 'normal' increased from study baseline for all dimensions, and there was a ≥ 46% reduction in the proportion of subjects with a rating of 'severe impairment'. CONCLUSION: These preliminary results suggest that escalating incobotulinumtoxinA doses up to 800 U are associated with improvement in quality of life ratings in subjects with multifocal upper- and lower-limb spasticity, and form a basis for future comparator studies. TRIAL REGISTRATION: ClinicalTrials.gov, NCT01603459. Date of registration: May 22, 2012.

Effect of Intrathecal Baclofen on Pain and Quality of Life in Poststroke Spasticity.

Background and Purpose- Intrathecal baclofen (ITB) is an effective treatment for managing patients with severe poststroke spasticity, who can experience continued pain and decline in their quality of life (QoL). SISTERS (Spasticity In Stroke-Randomized Study) was a randomized, controlled, open-label, multicenter, phase 4 study to evaluate ITB therapy versus conventional medical management (CMM) with oral antispastic medications for treatment of poststroke spasticity. Methods- Poststroke patients with spasticity in ≥2 extremities and an Ashworth Scale score of ≥3 in ≥2 affected lower extremity muscle groups were randomized (1:1) to ITB (N=31) or CMM (N=29). Both treatment arms received physiotherapy throughout. The primary outcome was the change in average Ashworth Scale score in the lower extremities of the affected side from baseline to month 6. Here, we report results for secondary outcomes: pain via the Numeric Pain Rating Scale, health-related QoL by the EuroQol-5 dimensional 3 level utility score and health status visual analog scale score, stroke-specific QoL, and patient satisfaction. Analyses were performed on an intention-to-treat basis. Results- We observed significant treatment effects in favor of ITB over CMM for changes from baseline to month 6 in Numeric Pain Rating Scale scores for actual pain (ITB versus CMM: mean, -1.17 [SD, 3.17] versus 0.00 [3.29]; median, -1.00 versus 0.00; P=0.0380) and least pain (mean, -1.61 [2.29] versus 0.24 [3.07]; median, -1.00 versus 0.00; P=0.0136), and EuroQol-5 dimensional 3 level utility scores (mean, +0.09 [0.26] versus +0.01 [0.16]; median, +0.07 versus 0.00; P=0.0197). Between-group differences were not statistically significant for EuroQol-5 dimensional 3 level visual analog scale, stroke-specific QoL summary, or Numeric Pain Rating Scale worst pain scores, although ITB patients showed greater numeric improvements from baseline during follow-up. More ITB patients than CMM patients (73% versus 48%) were satisfied with the spasticity reduction at month 6. Conclusions- These data support that ITB therapy is associated with improvements in pain and QoL in poststroke patients. Clinical Trial Registration- URL: https://www.clinicaltrials.gov . Unique identifier: NCT01032239.

Intrathecal baclofen therapy versus conventional medical management for severe poststroke spasticity: results from a multicentre, randomised, controlled, open-label trial (SISTERS).

BACKGROUND: Intrathecal baclofen (ITB) is a treatment option for patients with severe poststroke spasticity (PSS) who have not reached their therapy goal with other interventions. METHODS: 'Spasticity In Stroke-Randomised Study' (SISTERS) was a randomised, controlled, open-label, multicentre phase IV study to evaluate the efficacy and safety of ITB therapy versus conventional medical management (CMM) with oral antispastic medications for treatment of PSS. Patients with chronic stroke with spasticity in ≥2 extremities and an Ashworth Scale (AS) score ≥3 in at least two affected muscle groups in the lower extremities (LE) were randomised (1:1) to ITB or CMM. Both treatment arms received physiotherapy throughout. The primary outcome was the change in the average AS score in the LE of the affected body side from baseline to month 6. Analyses were performed for all patients as randomised (primary analysis) and all randomised patients as treated (safety analysis). RESULTS: Of 60 patients randomised to ITB (n=31) or CMM (n=29), 48 patients (24 per arm) completed the study. The primary analysis showed a significant effect of ITB therapy over CMM (mean AS score reduction, -0.99 (ITB) vs -0.43 (CMM); Hodges-Lehmann estimate, -0.667(95.1%CI -1.0000 to -0.1667); P=0.0140). More patients reported adverse events while receiving ITB (24/25 patients, 96%; 149 events) compared with CMM (22/35, 63%; 77 events), although events were generally consistent with the known safety profile of ITB therapy. CONCLUSIONS: These data support the use of ITB therapy as an alternative to CMM for treatment of generalised PSS in adults. TRIAL REGISTRATION NUMBER: NCT01032239; Results.

Development of progressive albuminuria in male Munich Wistar Frömter rats is androgen dependent.

Munich Wistar Frömter (MWF) rats develop spontaneous albuminuria that is linked to autosomal genetic loci and inherit a nephron deficit in both female and male animals, respectively. However, albuminuria and kidney damage are clearly more pronounced in males. Here we tested whether androgens and the androgen receptor influence albuminuria in male MWF. We first demonstrated in a pilot study that orchiectomy (Ox) of male MWF led to a significant suppression of urinary albumin excretion (UAE), while continuous testosterone supplementation in MWF Ox led to UAE levels similar to sham-operated (Sham) MWF rats. Subsequently, we performed a comparative main study between male MWF and normal Wistar rats to evaluate the effect of the androgen receptor on UAE development in adult animals up to the age of 18 wk. MWF Sham developed a marked increase in UAE compared with Wistar Sham (48.30 ± 6.16 vs. 0.42 ± 0.08 mg/24 h, P < 0.0001). UAE was significantly lower in MWF Ox compared with MWF Sham (-55%, P < 0.0001). In MWF Ox animals supplemented with testosterone and treated with the androgen receptor antagonist flutamide (OxTF) UAE at 18 wk was even lower compared with MWF Ox (-71%, P < 0.01) and similar to age-matched female MWF. The mRNA expression of renal tubular injury markers Kim1 and NGAL was increased in MWF Sham compared with Wistar Sham (P < 0.0008, respectively) and expression decreased significantly in MWF OxTF (P < 0.0004, respectively). Thus, the sexual dimorphism in albuminuria development in MWF can be attributed to testosterone and the androgen receptor in male rats.

Cardiorenal protection in experimental hypertension with renal failure: comparison between vasopeptidase inhibition and angiotensin receptor blockade.

OBJECTIVE: The aim of the present study was to compare the preventive impact of treatment with a vasopeptidase inhibitor (VPI) with an angiotensin-receptor blocker (ARB) on left ventricular (LV) function and renal damage in rats with renal failure after 5/6 renal ablation (Nx). METHODS: Rats (n = 15-20, each group) underwent either sham-operation (Sham) or 5/6 renal ablation (Nx). Two additional groups of Nx-animals (groups Nx-VPI and Nx-ARB) were treated with the VPI ilepatril (AVE7688, 30 mg kg(-1) d(-1)) or with the ARB olmesartan (10 mg kg(-1 )d(-1)). Animals were followed for 4 weeks. RESULTS: Systolic blood pressure (SBP), LV hypertrophy (LVH) and LV end-diastolic pressure (LVEDP) were increased 4 weeks after Nx (p < 0.05). LV pressure rise (+dP/dt/LVPmax), LV pressure fall (-dP/dt/LVPmax), and creatinine clearance decreased, while albuminuria and renal glomerulosclerosis index (GSI) increased with Nx (p < 0.05, respectively). In comparison to Nx, treatment with both VPI and ARB normalized SBP, LVH, LVEDP, +dP/dt/LVPmax, and -dP/dt/LVPmax to Sham control levels. GSI, but not creatinine clearance, was also normalized in response to both treatments. The significant increase in albuminuria observed in Nx (+230-fold versus Sham, p < 0.0001) was partially reduced in Nx-VPI (+47-fold versus Sham, p < 0.0001) and fully abolished in Nx-ARB. CONCLUSIONS: Both ilepatril and olmesartan conferred strong cardiorenal protective effects in rats with renal failure. While cardioprotection was clearly comparable with both treatment regimens, the ARB provided a better protection against the increase in albuminuria, although renal function and structural kidney changes were similarly affected by the VIP and ARB.

Genetic variants on rat chromosome 8 exhibit profound effects on hypertension severity and survival during nitric oxide inhibition in spontaneously hypertensive rats.

BACKGROUND: Hypertension and mortality is aggravated by nitric oxide inhibition with N(G)-nitro-L-arginine methyl ester (L-NAME) in spontaneously hypertensive rats (SHRs) but not in Munich Wistar Frömter (MWF) rats. MWF rats carry major albuminuria quantitative trait loci on rat chromosome (RNO) 6 and RNO8; susceptibility of SHRs to L-NAME is enhanced by transfer of RNO6 from MWF rats into the SHR background. Here, we tested whether the sensitivity to L-NAME in SHRs is affected by transfer of RNO8 from MWF rats in consomic SHR-8(MWF) rats. METHODS: In study 1, we analyzed survival in male SHR and SHR-8(MWF) rats in response to 18 weeks of treatment with either normal drinking water (vehicle-treated) or water containing 20mg/L L-NAME. In study 2, we analyzed blood pressure and renal damage in both strains in response to 6 weeks of treatment with L-NAME compared with vehicle-treated groups. RESULTS: In study 1, starting after 6 weeks of treatment with L-NAME, mortality reached 90% in SHRs in contrast with the group of L-NAME treated SHR-8(MWF) rats (P < 0.0001) in which all rats survived similar to vehicle-treated rats. In study 2, L-NAME resulted in a more pronounced increase in mean arterial blood pressures in SHRs compared with SHR-8(MWF) rats (216 ± 6 vs. 180 ± 11 mm Hg; P < 0.05). In contrast, tubulointerstitial kidney damage was even lower in SHRs compared with SHR-8(MWF) rats after L-NAME treatment (P < 0.05), whereas albuminuria was not different between strains. CONCLUSIONS: The blood pressure increase and impaired survival of SHRs in response to nitric oxide inhibition is profoundly influenced by genes on RNO8.

Induction of albuminuria and kidney damage in SHR by transfer of chromosome 8 from Munich Wistar Frömter rats.

Inbred Munich Wistar Frömter [MWF/FubRkb (RGD:724569), MWF] rats develop progressive albuminuria with age that is under polygenetic influence. We previously identified a major albuminuria quantitative trait locus (QTL) on rat chromosome (RNO)8 in MWF. To test the independent role of QTL(s) for albuminuria development on RNO8, we generated a consomic SHR-Chr 8(MWF)/Rkb (SHR-8(MWF)) strain by transferring RNO8 from MWF into the albuminuria-resistant background of the spontaneously hypertensive rat [SHR/FubRkb (RGD:631696; SHR)]. Young male MWF, SHR, and SHR-8(MWF) were sham-operated or unilaterally nephrectomized (Nx) at 6 wk and followed up to 24 wk of age, respectively. Systolic blood pressure was significantly lower in SHR-8(MWF) Sham compared with SHR Sham (-19.4 mmHg, P = 0.03) at 24 wk. In contrast, transfer of MWF-RNO8 into SHR induced a significant elevation of urinary albumin excretion (UAE) between weeks 12 and 24 in SHR-8(MWF) compared with SHR Sham animals (P < 0.0001, respectively). Nx resulted in a significant increase in UAE in both strains during follow-up (P < 0.0001, respectively), with significant higher values in SHR-8(MWF) compared with SHR (P < 0.005, respectively). Renal structural changes as determined by glomerulosclerosis (GSI) and tubulointerstitial damage index (TDI) were significantly higher in consomic animals either at Sham (TDI) or Nx (GSI) conditions (P < 0.05, respectively). These data confirm the independent role of MWF QTL(s) on RNO8 for both albuminuria and structural kidney damage. Moreover, this study shows for the first time the induction of albuminuria by transferring one or more albuminuria QTL into a resistant recipient background in a consomic rat strain.

Elimination of severe albuminuria in aging hypertensive rats by exchange of 2 chromosomes in double-consomic rats.

The inherited nephron deficit and progressive albuminuria development observed in hypertensive Munich Wistar Frömter (MWF) rats are influenced by quantitative trait loci on rat chromosome (RNO) 6 and RNO8. Previous studies in young MWF rats suggested that the nephron deficit represents a cause for glomerular hypertrophy preceding onset of albuminuria at 8 weeks and demonstrated a simultaneous induction of the podocyte stress marker desmin and podoplanin loss in podocytes. Here we investigated the separate genetic influence of RNO6 and RNO8 on early glomerular changes and subsequent albuminuria in single-consomic MWF rats in which RNO6 (MWF-6(SHR)) and RNO8 (MWF-8(SHR)) were replaced by the respective spontaneously hypertensive rat (SHR) chromosome. Furthermore, we tested the role of synergistic effects between both chromosomes in a double-consomic MWF-6(SHR)8(SHR) strain. Increased glomerular, extramesangial desmin expressions at 6 and albuminuria at 8 weeks were significantly reduced in single- and double-consomics (P<0.05 versus MWF, respectively). MWF-6(SHR)8(SHR) rats demonstrated the lowest desmin expression and glomerular volume (P<0.05 versus MWF, MWF-6(SHR), and MWF-8(SHR), respectively), indicating synergistic effects between RNO6 and RNO8. A significant and similar loss of podoplanin was only seen in MWF and MWF-6(SHR) rats but not in MWF-8(SHR) and MWF-6(SHR)8(SHR) rats (P<0.02, respectively); this refutes a mandatory coupling of desmin induction and podoplanin loss in podocytes preceding albuminuria and reveals a genetic link between RNO8 and loss of podoplanin protein. Long-term follow up in MWF-6(SHR)8(SHR) rats demonstrates the relevance of the absence of glomerular changes in young animals, because double-consomics demonstrate a complete suppression of progressive albuminuria and kidney damage compared with MWF rats despite similar blood pressures.

Genetic locus on MWF rat chromosome 6 affects kidney damage in response to L-NAME treatment in spontaneously hypertensive rats.

A major quantitative trait locus (QTL) on rat chromosome (RNO)6 was linked to albuminuria in Munich Wistar Frömter rats (MWF). We tested whether transfer of MWF RNO6 into the background of albuminuria-resistant spontaneously hypertensive rats (SHR) induces albuminuria in consomic SHR-6(MWF) animals. Male MWF, SHR, and SHR-6(MWF) were sham operated and treated between 6 and 24 wk of age with normal water (Sham) or with water containing 20 mg/l N(G)-nitro-L-arginine methyl ester (L-NAME) or unilaterally nephrectomized (Nx). Compared with SHR albuminuria was not increased in SHR-6(MWF) in both Sham and Nx groups. All animals survived the observation period in Sham and Nx groups, while premature mortality occurred from 12-14 wk on in L-NAME-treated SHR and SHR-6(MWF) compared with MWF L-NAME animals, in which survival was not affected (P < 0.005, respectively). Subsequent further analysis of L-NAME-treated animals at 12 wk of age showed significantly increased arterial blood pressures in both SHR and SHR-6(MWF) compared with control (P < 0.05), with higher levels in SHR compared with consomics (P < 0.05). However, L-NAME-treated consomic animals demonstrated increased albuminuria compared with SHR (12.7 +/- 3.5 vs. 0.8 +/- 0.2 mg/24 h; P < 0.05) and an induction of tubulointerstitial structural injury and expression of neutrophil gelatinase-associated lipocalin mRNA (P < 0.05 vs. other strains). Our study demonstrates that isolation of the RNO6 albuminuria QTL from the MWF background and transfer into SHR fails to induce an albuminuria phenotype during normal conditions or after nephron reduction. Moreover, our data indicate that genes on RNO6 contribute to the development of L-NAME-induced renal damage in the SHR strain.

Characterization of human chondrocytes exposed to simulated microgravity.

BACKGROUND: Tissue engineering is a strategy of cartilage regeneration, but scaffolds, required for 3D growth of chondrocytes, are still a problem. METHODS: Searching for possibilities to improve scaffold-free engineering of cartilage, we characterized human chondrocytes incubated on a random positioning machine (RPM) to simulate microgravity (microg). RESULTS: When cultured in simulated microg, human chondrocytes start forming 3D cell assemblies within 5 days. After 24h, we could not detect caspase-3, Fas, p53 or Bcl-2 proteins in these cells, Annexin V flow cytometry, however, revealed 18% of apoptotic chondrocytes in 1g cultures but only 10% on the RPM. Both rates of apoptosis were not changed, when vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF) was added. 24 h, simulated microgravity also had significantly decreased collagen type I and X, but did not change collagen type IV and laminin, while collagen type II, chondroitin sulfate and aggrecan were elevated as compared with 1g controls. The production of collagen type II/X, chondroitin sulfate and aggrecan was modified, when external bFGF or VEGF had been applied. CONCLUSION: Chondrocytes exposed to simulated microg seem to change their extracellular matrix production behavior, while they rearrange their cytoskeletal proteins prior to forming 3D aggregates.

Short-term treatment with a beta-blocker with vasodilative capacities improves intrarenal endothelial function in experimental renal failure.

AIMS: In patients with renal disease the cardiovascular risk is greatly increased, and endothelial dysfunction is assumed to play a pivotal role in this process. Therefore we compared treatment effects of a beta-blocker with additional vasodilatory capacities (nebivolol) and a beta-blocker lacking these actions (metoprolol) on intrarenal and coronary vascular function in a rat model of renal failure with hypertension. MAIN METHODS: Renal failure was induced by 5/6-nephrectomy (Nx) and analyzed after 4 weeks in Wistar rats. Untreated Nx, Nx/nebivolol 6 mg/d (Nx-Nebi); Nx/metoprolol 60 mg/d (Nx-Meto) and sham-operated (Sham) animals were studied. Isolated small renal and coronary arteries were investigated for endothelium-dependent relaxation to acetylcholine (ACh) and for the contribution of the endothelial mediators NO and endothelium-derived hyperpolarizing factor (EDHF). KEY FINDINGS: Systolic blood pressure (SBP) was significantly increased in Nx, Nx-Nebi, and Nx-Meto (168+/-5, 153+/-3, and 162+/-6 mmHg) compared to Sham (138+/-3 mmHg, p<0.05, respectively). The increase in albuminuria of Nx (120-fold vs. Sham, p<0.0001) was almost (-85%) normalized by nebivolol compared to Sham (p<0.05), whereas metoprolol induced no significant effect. Renal arteries showed significantly increased Ach-relaxation in Nx and Nx-Nebi (Emax 86+/-4% and 76+/-7%, p<0.05) due to an increase in EDHF-mediated dilation (Emax_EDHF 78+/-7% and 73+/-6%) compared to Sham (Emax 54+/-4% and Emax_EDHF 44+/-6%) and Nx-Meto (Emax 42+/-12% and Emax_EDHF 18+/-5%). ACh-relaxation in coronary arteries was similar between groups but the contribution of NO (relative to EDHF) was strongly increased by nebivolol. SIGNIFICANCE: The present findings offer an explanation of the nephroprotective effect of intrarenal endothelial function in renal failure.

A delayed type of three-dimensional growth of human endothelial cells under simulated weightlessness.

Endothelial cells (ECs) form three-dimensional (3D) aggregates without any scaffold when they are exposed to microgravity simulated by a random positioning machine (RPM) but not under static conditions at gravity. Here we describe a delayed type of formation of 3D structures of ECs that was initiated when ECs cultured on a desktop RPM remained adherent for the first 5 days but spread over neighboring adherent cells, forming little colonies. After 2 weeks, tube-like structures (TSs) became visible in these cultures. They included a lumen, and they elongated during another 2 weeks of culturing. The walls of these TSs consisted mainly of single-layered ECs, which had produced significantly more beta(1)-integrin, laminin, fibronectin, and alpha-tubulin than ECs simultaneously grown adhering to the culture dishes under microgravity or normal gravity. The amount of actin protein was similar in ECs incorporated in TSs and in ECs growing at gravity. The ratio of tissue inhibitor of metalloproteinases-1 to matrix metalloproteinase-2 found in the supernatants was lower at the seventh than at the 28th day of culturing. These results suggest that culturing ECs under conditions of modeled gravitational unloading represents a new technique for studying the formation of tubes that resemble vascular intimas.

Genetic analysis of salt-sensitive hypertension in Dahl rats reveals a link between cardiac fibrosis and high cholesterol.

AIMS: Previously we confirmed an important role of rat chromosome 19 (RNO19) for salt-sensitive hypertension and target organ damage in male Dahl salt-sensitive rats (SS rats). The aim of this study was to further analyse the basis of left ventricular (LV) fibrosis development in both male and female rats in this model. To this end we utilized a consomic SS-19(SHR) rat strain in which RNO19 was transferred from spontaneously hypertensive rats (SHR) into the susceptible background of SS. METHODS AND RESULTS: We compared the effects of low- (0.2% NaCl) and high-salt (4% NaCl) diet on the development of hypertension, blood lipids and LV fibrosis in male and female SS, SHR, and SS-19(SHR) rats. Systolic blood pressure was significantly lower in male and female SS-19(SHR) compared with SS under both diets (P < 0.001). Relative LV weight was similarly reduced in SS-19(SHR) compared with SS in either sex. Plasma cholesterol concentrations were significantly elevated in high-salt fed male and female SS (141 +/- 6 and 110 +/- 7 mg/dL) compared with SHR (47 +/- 2 and 62 +/- 8 mg/dL, P < 0.001) and were significantly lowered in male and female consomic rats (100 +/- 7 and 87 +/- 3 mg/dL). Both LV interstitial fibrosis (LVIF) and perivascular fibrosis (LVPF) were significantly reduced in high-salt male and female SS-19(SHR). A significant correlation between cholesterol concentrations and LVPF (r = 0.464) and LVIF (r = 0.401, P < 0.0001, respectively) was detected. Fibrosis parameters demonstrated no correlation with blood pressure, LV weight or plasma triglycerides concentrations. LV immunohistochemistry analysis showed a significant higher number of ED-1 positive cells in SS compared with SS-19(SHR). Depositions of collagen I and fibronectin were also greater in LV tissue of SS compared with SS-19(SHR). CONCLUSION: Our findings point to a link between hypercholesterolemia and LV fibrosis in salt-sensitive hypertension of SS rats which is genetically modulated by RNO19.

Nephron deficit is not required for progressive proteinuria development in the Munich Wistar Frömter rat.

The Munich Wistar Frömter (MWF) rat represents a genetic model with an inherited nephron deficit and exhibits mild hypertension and progressive albuminuria, which is more pronounced in males than females. Previously, we demonstrated in a consomic strain that replacement of a quantitative trait locus on chromosome 6 normalized the nephron deficit and suppressed albuminuria development, suggesting a link between the two findings. Here we tested the role of a second major locus linked to albuminuria in MWF on chromosome 8 and generated the consomic strain MWF-8(SHR) by transfer of chromosome 8 from spontaneously hypertensive rats (SHR) into MWF. The early onset of albuminuria at 8 wk of age in MWF (>50-fold increase compared with SHR) was significantly suppressed in consomic animals, and the development of marked proteinuria at 32 wk significantly diminished. Total nephron number in consomic rats (23,771 +/- 1,352) and MWF (27,028 +/- 1,322) were similar and significantly lower (-36%) compared with SHR (36,979 +/- 1,352, P < 0.0001). The development of mild albuminuria in female MWF was also significantly diminished in MWF-8(SHR). Thus, the development of overt and mild albuminuria in male and female MWF rats is not a mandatory consequence of the inherited nephron deficit. The locus on chromosome 8 appears of interest, because its exchange between MWF and SHR protects against the development of albuminuria in MWF-8(SHR) animals despite their inherited nephron deficit and higher systolic blood pressure.

Protective effect of female gender on the development of albuminuria in a polygenetic rat model is enhanced further by replacement of a major autosomal QTL.

Clinical and experimental studies indicate that the progression of renal disease is faster in males than females. These observations are corroborated by a sexual dimorphism observed in the polygenetic MWF (Munich Wistar Frömter) rat model. The age-dependent spontaneous progression of increased UAE (urinary albumin excretion) in male MWF rats is influenced by multiple QTLs (quantitative trait loci). In contrast, female MWF rats only develop a slight increase in UAE, while the role of genetic factors for this phenotype is unknown. In the present study, we show that, compared with resistant SHRs (spontaneously hypertensive rats), both male and female MWF rats develop a significant increase in UAE at 24 weeks of age (P<0.0001), although blood pressures were lower compared with SHRs (P<0.0001). UAE was significantly higher in male (7-fold) compared with female MWF rats (162.6+/-15.9 compared with 24.0+/-5.5 mg/24 h respectively; P<0.0001), and only male MWF rats developed significant glomerulosclerosis and tubulointerstitial damage in the kidney (P<0.0001). To test the role of genetic factors in the development of low grade albuminuria in female MWF rats, we analysed the role of a major UAE QTL on rat chromosome 6. To this end, we analysed a consomic MWF-6(SHR) strain in which chromosome 6 from SHRs was introgressed into the MWF rat background. Time course analysis of UAE in females indicated that the small increase in UAE in MWF rats was fully suppressed by exchange of rat chromosome 6. Thus, taken together with previous studies in males, we show that RNO6 protects against the increase in albuminuria with age in both female and male MWF rats.

Expression of vascular endothelial growth factor and receptor tyrosine kinases in cardiac ischemia/reperfusion injury.

INTRODUCTION: Vascular endothelial growth factor (VEGF) expression is regulated by hypoxia and cytokines, including insulin-like growth factor (IGF)-1. We examined the influence of ischemia/reperfusion (I/R) on IGF-1, VEGF, fetal liver kinase (flk-1), fms-like tyrosine kinase-1 (flt-1), and laminin using an isolated hemoperfused working porcine heart model of acute ischemia (2 h) and reperfusion (4 h). METHODS: Twenty-three porcine hearts were randomized into the following groups: five nonischemic control hearts (Group C), five I/R hearts with occlusion of the ramus circumflexus; three I/R hearts treated with quinaprilat, a potent angiotensin-converting enzyme (ACE) inhibitor (Group Q); five I/R hearts treated with angiotensin I (Group Ang I), and 5 I/R hearts treated with Ang I and quinaprilat (Group QA). RESULTS: Compared to C, VEGF mRNA and protein contents were significantly increased in I/R and Ang I hearts. flk-1 and flt-1 were increased in I/R (2.2-/1.95-fold) and further elevated by Ang I (3.2-/3.4-fold) compared with C. Quinaprilat application attenuated the amount of VEGF significantly and of flk-1 slightly but not that of flt-1. In contrast, IGF-1 and IGF-1 receptor (IGF-1R) proteins were elevated in I/R hearts (3-/1.4-fold vs. C) and further increased in the presence of Q. These findings were accompanied by an elevation of laminin mRNA and protein levels. Moreover, we observed an increase in collagen Type IV and chondroitin sulfate content in I/R (2.9-/1.4-fold) and Ang I (3.5-/1.5-fold) hearts. Quinaprilat significantly reduced laminin and chondroitin sulfate proteins. CONCLUSION: These data suggest that the VEGF/VEGF receptor and IGF-1-IGF-1R systems are activated by I/R. The benefits of ACE inhibition in attenuation of cardiac remodeling may be mediated by IGF-1.

Development of overt proteinuria in the Munich Wistar Frömter rat is suppressed by replacement of chromosome 6 in a consomic rat strain.

In a cross between the Munich Wistar Frömter (MWF) rat and spontaneously hypertensive rats (SHR), a major quantitative trait locus (QTL) was identified on rat chromosome 6 (RNO6) that demonstrated the strongest linkage to albuminuria among several QTL identified. The QTL represented the only locus that is linked to both early-onset albuminuria and increased renal interstitial fibrosis in adult animals. A consomic MWF-6(SHR) strain in which chromosome 6 from SHR was introgressed into the MWF background therefore was generated to test the relevance of this QTL. Phenotype analysis at 8 wk of age revealed that early onset of albuminuria in MWF with a 55-fold elevation of urinary albumin excretion compared with SHR (P < 0.0001) was completely abolished in MWF-6(SHR). Time-course analysis until week 24 demonstrated only a moderate increase of urinary albumin excretion in MWF-6(SHR), whereas MWF reached levels in the nephrotic range (16.6 +/- 3.5 versus 162.6 +/- 16.0 mg/24 h; P < 0.0001). At this age, analysis of glomerulosclerosis, tubulointerstitial damage, renal interstitial fibrosis, and renal collagen III mRNA expression revealed a significant improvement of all parameters in MWF-6(SHR) compared with MWF (P < 0.05). At 32 wk, MWF but not MWF-6(SHR) demonstrated overt proteinuria (354.6 +/- 37.6 versus 48.8 +/- 13.2; P < 0.0001), whereas serum urea, cholesterol, and triglyceride concentrations were lower and creatinine clearance was higher in MWF-6(SHR) compared with MWF (P < 0.05). Therefore, although albuminuria in MWF is determined by a complex interplay of several QTL, our data demonstrate that genetic exchange of one locus on RNO6 leads to marked suppression of early-onset albuminuria and renal damage in MWF.

Rat chromosome 19 transfer from SHR ameliorates hypertension, salt-sensitivity, cardiovascular and renal organ damage in salt-sensitive Dahl rats.

OBJECTIVES: Unlike Dahl salt-sensitive (SS) rats, some strains of spontaneously hypertensive (SHR) rats develop only minor organ damage even when exposed to high-salt diet. In previous linkage studies, we identified quantitative trait loci on rat chromosome 19 (RNO19) linked to the SHR allele suggesting a protective effect against salt-induced hypertensive organ damage in SS. METHODS: To test the relevance of this finding, we generated and characterized a consomic strain SS-19SHR in which RNO19 from SHR was introgressed into the susceptible background of SS. We compared the effects of low-salt (0.2% NaCl) and high-salt (4% NaCl) diet exposure for 8 weeks on the development of hypertension and target organ damage in male consomic and SS animals (n=14-20, each). RESULTS: Systolic blood pressure, relative left ventricular weight and urinary protein excretion were significantly lower in SS-19SHR compared to SS under both low-salt and high-salt diet (P < 0.05, respectively). Left ventricular atrial natriuretic peptide mRNA expression showed a more pronounced 4.5-fold increase in SS compared to SS-19 (two-fold) after high-salt (P < 0.05). In comparison to low diet, high-salt exposure induced a significant increase in vascular aortic hypertrophy index, left ventricular interstitial fibrosis (+210%) and perivascular fibrosis (+195%) in SS but not in consomic SS-19SHR (P < 0.05, respectively). CONCLUSIONS: These results demonstrate a strong protective effect of RNO19 from SHR on the development of hypertension, salt-sensitivity, cardiovascular and renal organ damage in SS. In particular, we demonstrate a genetic effect protecting against the development of cardiac fibrosis in salt-sensitive hypertension.