Differentiation of bone marrow-derived cells into regenerated mesothelial cells in peritoneal remodeling using a peritoneal fibrosis mouse model.

Marked thickening of the peritoneum and vasculopathy in the submesothelial compact zone have been reported in long-term peritoneal dialysis patients. Bone marrow (BM)-derived cell lines are considered to be useful tools for therapy of various diseases. To clarify the role of BM-derived cells in the peritoneal fibrosis (PF) model, we analyzed several lineages of cells in the peritoneum. BM cells from green fluorescent protein (GFP) transgenic mice were transplanted into naïve C57Bl/6 mice. Chlorhexidine gluconate (CG) was injected intraperitoneally to induce PF. Immunohistochemical analysis was performed with parietal peritoneum using anti-Sca-1 or -c-Kit and -GFP antibodies. Isolated BM cells were also transplanted into the CG-stimulated peritoneum. BM-derived cells from GFP transgenic mice appeared in the submesothelium from days 14 to 42. Both GFP- and stem cell marker-positive cells were observed in the submesothelium and on the surface. Isolated c-Kit-positive cells, transplanted into the peritoneal cavity, differentiated into mesothelial cells. In this study, we investigated whether or not BM-derived cells play a role in the repair of PF and immature cells have the potential of inducing repair of the peritoneum. The findings of this study suggest a new concept for therapy of PF.

Cardiac function and structure in longitudinal analysis of echocardiography in peritoneal dialysis patients.

BACKGROUND: Echocardiography is widely used for the evaluation of cardiac structures and function. The prognostic value of assessment of left cardiac atrium (LA) size in peritoneal dialysis (PD) patients is still unclear. The objective of the present study is to investigate prospectively a longitudinal monitoring of echocardiography parameters after start of PD. We also investigated a correlation study among plasma atrial natriuretic peptide (ANP) level, LA size, and cardiac function undergoing aggressive treatment. METHODS: Correlation among plasma ANP, LA size, and cardiac function was prospectively analyzed by Doppler echocardiography in 32 PD patients in Juntendo University Hospital, Tokyo. Measurement of these parameters was performed at 0, 6, 12, 18, and 24 months after start of PD. All patients were treated with an angiotensin type 1 receptor blocker to control blood pressure to less than 140/90 mmHg. Other antihypertensive drugs such as diuretics and/or calcium channel blockers were added if blood pressure rose to over 140/90 mmHg. Hemoglobin and hematocrit levels were targeted at 10.0 g/dL and 30.0% respectively with recombinant human erythropoietin treatment. A diuretic was added or patients decreased their water intake if ANP was more than 43.0 pg/mL or LA diameter (LAD) more than 39 mm, and for other basic markers of volume status. Cardiac function was measured before and after drainage of PD fluid to evaluate the influence of cardiac function. RESULTS: LAD at start of dialysis (36 +/- 4.6 mm) decreased significantly to 33 +/- 3.3 mm (p < 0.05), 33 +/- 3.2 mm (p < 0.05), and 33 +/- 3.6 mm (p < 0.05) after 6, 12, and 24 months, respectively. Ejection fraction after 6 months was significantly increased compared with that at start of dialysis (p < 0.05). Left ventricular mass index (LVMI) after 6, 12, and 24 months was significantly decreased compared with that at start of dialysis (p < 0.05). ANP was 56 +/- 39 pg/mL at start of dialysis and decreased significantly to 33 +/- 19 pg/mL after 24 months (p < 0.05). ANP was significantly correlated with LAD (r = 0.412, p < 0.01), transmitral A wave flow velocity (r = 0.429, p < 0.01), and LVMI (r = 0.426, p < 0.01). Instillation of the dialysis fluid did not affect any parameters except inferior vena cava dimension. CONCLUSION: This study demonstrates a reduction in LA size and LVMI in PD patients followed over 24 months. Left ventricular structure, contraction, and compliance were well preserved in PD patients undergoing aggressive treatment based on measurements of plasma ANP and LAD.

Inhibitory effects of matrix metalloproteinase inhibitor ONO-4817 on morphological alterations in chlorhexidine gluconate-induced peritoneal sclerosis rats.

BACKGROUND: The activity of gelatinase, matrix metalloproteinase-2, in effluent was increased in peritoneal dialysis patients with encapsulated peritoneal sclerosis (EPS) and in chlorhexidine gluconate-induced peritoneal sclerosing (PS) animal models. The objective of the present study was to investigate the effect of matrix metalloproteinase inhibitor (ONO-4817), an anticancer agent with anti-angiogenesis and anti-infiltration effects, on the development of peritoneal fibrosis in chlorhexidine gluconate-induced PS rats. METHODS: Forty-five Sprague-Dawley (S-D) rats were intraperitoneally injected with saline as control (n = 15) or with chlorhexidine gluconate (CH) (1.5 ml/100 g) in the CH group (n = 15). ONO-4817 (5 mg/rat) was administered intravenously to CH rats (the ONO-4817 group, n = 15) from initiation to the end of the study. After 22 days of ONO-4817 administration, the rats were sacrificed and the parietal peritoneum was harvested. The gene expressions of transforming growth factor-beta (TGF-beta), alpha-smooth muscle actin (alpha-SMA) and type I collagen in the peritoneum were analysed by the reverse transcription-polymerase chain reaction (RT-PCR). Peritoneal tissues were also evaluated immunohistologically. RESULTS: ONO-4817 significantly inhibited thickening of the submesothelial layer and accumulation of type I collagen in the peritoneum. ONO-4817 also prevented increases of the number of macrophages and blood vessels. The expressions of TGF-beta, alpha-SMA and type I collagen in the peritoneum were markedly suppressed in ONO-4817-treated rats. CONCLUSION: It appears that the administration of the MMP inhibitor ONO-4817 might be a new approach to the amelioration of PS.

Pharmacokinetics of calcitriol and maxacalcitol administered into peritoneal dialysate bags in peritoneal dialysis patients.

OBJECTIVES: It is well known that injection of calcitriol (CT) or maxacalcitol (OCT) is very effective in hemodialysis patients with secondary hyperparathyroidism (2HPT). However, it is difficult to use these drugs with peritoneal dialysis (PD) patients with 2HPT because these drugs must be injected two or three times per week. The objective of the present study was to evaluate the stability of physiological activities of CT and OCT in PD bags and to determine the CT or OCT dosage for intraperitoneal (IP) administration. MATERIALS AND METHODS: We added CT 1.5 microg or OCT 10 microg to Dianeal PD-2 (approximate pH = 5.0, calcium = 0.87 mmol/L; Baxter,Tokyo, Japan), Midpeliq 250 (approximate pH = 7.0, Ca = 1.0 mmol/L;Terumo Corporation, Tokyo, Japan), and Peritoliq 250 (approximate pH = 5.5, Ca = 1.0 mmol/L; Terumo Corp.). Dialysis solutions were collected from the PD bags at 0, 1, 4, 8, 12, 24, 48, and 72 hours after addition of CT and OCT. The activities of CT and OCT in the dialysis effluent were measured by radioimmunoassay. The levels of serum and effluent OCT after a single IP administration of 10 microg OCT were examined in 4 PO patients with advanced 2HPT. RESULTS: Although the levels of CT and OCT in PD bags made of polyvinyl resins decreased by 70% - 75% immediately after injection, levels in PD bags made of polypropylene resins decreased only slightly. The concentration of CT mixed into the acidic solution in glass containers was stable; the decreased concentration of CT in the PD solution might be due to adsorption onto polyvinyl resins. The maximum serum concentration after IP administration of 10 microg OCT was 750 pg/mL after 5 minutes, and remained at 500 pg/mL at 60 minutes. These results show good peritoneal transport of OCT but not rapid disappearance, unlike intravenous administration. CONCLUSIONS: If peritoneal administration of vitamin D derivatives is contemplated, it is important to select the composition of PD bag resins, type of vitamin D analog, and time lag to use when deciding the dosage of injectable vitamin D preparations, such as OCT or CT, for IP administration to PD patients. It appears that IP administration in overnight dwells might be useful for PD patients as a complementary vitamin D preparation.

Rapid, simple, and reliable method for the diagnosis of CAPD peritonitis using the new MMP-9 test kit.

The objective of the present study was to evaluate the sensitivity and efficiency of the matrix metalloproteinase-9 (MMP-9) test kit for the diagnosis of bacterial peritonitis in patients undergoing peritoneal dialysis (PD). Peritoneal effluents were collected from seven continuous ambulatory PD (CAPD) patients with peritonitis, four patients with suspected peritonitis, 30 maintenance PD patients without infection, and seven patients at initiation of PD. The MMP-9 test kit was used to analyze 112 peritoneal effluent samples. These peritoneal effluents were also used to count leukocytes and examine microorganisms. MMP expression was measured by gelatin zymography, and activities were measured by an enzyme-linked immunosorbent assay (ELISA). The relationship between the reactivity of the test kit and the number of leukocytes in the samples was examined. There was a significant difference in the number of leukocytes in peritoneal effluents between the negative and positive groups detected by the MMP-9 test kit (P < 0.0001). The results obtained with the MMP-9 test kit were negative for peritoneal effluent samples that did not show increased cell counts. The reactivity of the MMP-9 test kit showed no significant differences among various microorganisms, and remained stable. The MMP-9 test kit appears to be a simple and reliable method for early diagnosis of CAPD peritonitis, and reflects the leukocyte count in peritoneal effluents.

Morphologic changes of peritoneum and expression of VEGF in encapsulated peritoneal sclerosis rat models.

BACKGROUND: Simple sclerosis consisted of a thin layer of submesothelial sclerotic tissue in peritoneal dialysis patients. Encapsulated peritoneal sclerosis (EPS) was characterized by thick sclerotic tissue involving vascular alterations in peritoneal dialysis patients. The objective of the present study is to evaluate serial morphologic changes and expressions of angiogenic factors [i.e., vascular endothelial growth factor (VEGF), angiopietin-1 (Ang-1), and angiopoietin-2 (Ang-2)] in EPS rat models. METHODS: Twenty-four rats were given a daily intraperitoneal injection of chlorhexidine gluconate and ethanol dissolved in saline (CH). Nine rats were injected with CH and anti-VEGF neutralizing antibody simultaneously. Quantitative blood vessel evaluation was performed by staining for GS1-lectin. The mRNA expression of VEGF, Ang-1, Ang-2, and their receptors was evaluated by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Immunohistochemical staining was performed in peritoneal vessels using anti-VEGF, Ang-1, and Ang-2 antibodies. Hematopoietic stem cells were detected using anti-CD34 antibody. RESULTS: The vessel area, diameter, and length gradually increased until day 21, and then decreased. VEGF and Ang-2 mRNA expressions gradually increased until day 35. In contrast, Ang-1 peaked at day 21 and then decreased significantly. VEGF blockade improved the experimental EPS. In immunohistochemistry, the vessels stained by VEGF and Ang-2 were detected in subfibrous layer. CD34-positive cells were markedly stained at day 21. CONCLUSION: Neoangiogenesis was observed in a rat model of experimental EPS. VEGF and angiopoietin/Tie system play an important role in the neoangiogenesis in this model. An analysis using this experimental rat model may elucidate the development of EPS in peritoneal dialysis patients.