Alendronate does not prevent long bone fragility in an inactive rat model

The lack of estrogen and inactivity are both important in the pathogenesis of osteoporosis in elderly women, and there have been no appropriate rodent studies to examine the effects of common bisphosphonates on these two components separately. We compared the efficacy of alendronate (ALN) on the long bones of aged female rats, which were sedentary, estrogen deficient, or both. The rats were either forced to remain in a sitting position or allowed to walk in standard cages with or without ALN administration. The 8-week experimental period began 5 weeks after ovariectomy or sham surgery. Parameters of the hindlimb bones were determined by a three-point bending test, peripheral quantitative computed tomography, microfocus computed tomography, confocal laser Raman microspectroscopy, and dynamic histomorphometry. Regardless of ovariectomy, ALN was ineffective against the deterioration of breaking stress caused by sitting even though the trabecular bone mineral density was significantly higher in the sitting–ALN groups. Toughness was significantly deficient in the ovariectomy sitting–ALN group. This was in agreement with the bone geometry with a greater marrow space. Sitting also increased the mineral-to-matrix ratio and the carbonate-to-phosphate ratio, both indicative of aged bone. A greater loss of proteinaceous amide intensity compared with mineral intensity resulted in an increased mineral-to-matrix ratio in the presence of ALN. Sitting resulted in deficits in the quality and the geometry of cortical bone, resulting in fragility. The use of bisphosphonates, such as ALN, may provide a therapy best suited for osteoporotic individuals whose daily activity is not limited.     

Impact of turbulent blood flow in the aortic root on de novo aortic insufficiency during continuous-flow left ventricular-assist device support

Severe aortic insufficiency (AI) after implantation of continuous-flow left ventricular-assist device (LVAD) affects device performance and outcomes. However, the mechanism for the occurrence and progression of AI has not been elucidated. We investigated the impact of nonphysiological retrograde blood flow in the aortic root on AI after LVAD implantation. Blood flow pattern was analyzed in patients with and without AI (n = 3 each) who underwent LVAD implantation, by computational fluid dynamics with patient-specific geometries, which were reproduced using electrocardiogram-gated 320-slice computed tomographic images. The total volume of retrograde blood flow during one cardiac cycle (716 ± 88 mL) was higher and the volume of slow blood flow (<0.1 cm/s) (0.16 ± 0.04 cm³) was lower in patients with AI than in those without AI (360 ± 111 mL, P = .0495, and 0.49 ± 0.08 cm³, P = .0495, respectively). No significant difference in wall shear stress on the aortic valve was observed between the groups. Patients with AI had a perpendicular anastomosis at the distal ascending aorta and the simulation in the modified anastomosis model of patients with AI showed that the retrograde blood flow pattern depended on the angle and position of anastomosis. Computational fluid dynamics revealed strong retrograde blood flow in the ascending aorta and aortic root in patients with AI after LVAD implantation. The angle and position of LVAD outflow anastomosis might impact retrograde blood flow and de novo AI after LVAD implantation.     

Very Short-Term Effects of the Dipeptidyl Peptidase-4 Inhibitor Sitagliptin on the Secretion of Insulin,Glucagon, and Incretin Hormones in Japanese Patients with Type?2 Diabetes Mellitus: Analysis of Meal Tolerance Test Data

Background

Sitagliptin inhibits dipeptidyl peptidase-4, which inactivates the incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide. To assess its antidiabetic potency, we used meal tolerance tests (MTTs) to determine the very short-term effects of sitagliptin on plasma concentrations of insulin and glucagon.

Methods

On day 1, patients with newly diagnosed or uncontrolled type 2 diabetes mellitus started a calorie-restricted diet. On day 2, the first MTT was performed, before treatment with sitagliptin 50 mg/day started later the same day. On day 5, a second MTT was performed. Area under the concentration–time curves (AUCs) of relevant laboratory values were calculated [AUC from time zero to 2 h (AUC_0–2h) and from time zero to 4 h (AUC_0–4h)].

Results

Fifteen patients were enrolled. AUCs for postprandial plasma glucose were decreased after 3 days of sitagliptin treatment [AUC_0–2h 457 ± 115 mg/dL·h (25.4 ± 6.4 mmol/L·h) to 369 ± 108 mg/dL·h (20.5 ± 6.0 mmol/L·h); AUC_0–4h 896 ± 248 mg/dL·h (49.7 ± 13.8 mmol/L·h) to 701 ± 246 mg/dL·h (38.9 ± 13.7 mmol/L·h); both p < 0.001]. AUC_0–2h and AUC_0–4h for postprandial plasma glucagon also decreased: 195 ± 57 to 180 ± 57 pg/mL·h (p < 0.05) and 376 ± 105 to 349 ± 105 pg/mL·h (p < 0.01), respectively. The AUC_0–2h [median with quartile values (25 %, 75 %)] for active GLP-1 increased: 10.5 (8.5, 15.2) to 26.4 (16.7, 32.4) pmol/L·h (p = 0.03).

Conclusions

Very short-term (3-day) treatment with sitagliptin decreases postprandial plasma glucose significantly. This early reduction in glucose may result partly from suppression of excessive glucagon secretion, through a direct effect on active GLP-1. Improvement in postprandial plasma glucose, through suppression of glucagon secretion, is believed to be an advantage of sitagliptin for the treatment of patients with type 2 diabetes.     

Optimal Initial Dose of Orally Administered Once-daily Extended-release Tacrolimus Following Intravenous Tacrolimus Therapy After Liver Transplantation

Introduction

Once-daily extended-release tacrolimus (Tac-OD) is expected to reduce non-adherence in recipients after liver transplantation (LT). The aim of this study was to determine the optimal initial dose of orally administered Tac-OD after intravenous tacrolimus (Tac-IV) therapy after LT.

Patients and Methods

This prospective study included 10 adult recipients who had undergone LT at our institute. The recipients were prescribed tacrolimus by continuous intravenous administration with a steroid as initial immunosuppression therapy. Tacrolimus was converted from intravenous administration to once-daily oral intake when gastrointestinal function returned. We evaluated tacrolimus concentrations in blood 9 times a day and area under the blood concentration−time curve (AUC) during conversion. The optimal initial dose of Tac-OD was determined based on simple regression analysis between the oral dose of Tac-OD and the total dose of Tac-IV during a 24-hour period.

Results

The AUC before and after conversion showed no differences. We found that the optimal initial dose of Tac-OD was 8 times the dose of Tac-IV. There was a relationship between the AUC and the trough level. No recipients experienced acute rejection or adverse effects such as renal failure, neurotoxicity, or cardiac failure during conversion.

Conclusions

We successfully converted continuous Tac-IV to oral intake of Tac-OD by adjusting the dose using trough levels without acute rejection or adverse effects. The AUC of Tac-OD correlated with the trough level. The optimal initial dose ratio of Tac-OD after Tac-IV was 8:1.     

Regulation of Macrophage-Mediated Xenocytotoxicity by Overexpression of Alpha-2,6-sialyltransferase in Swine Endothelial Cells

Macrophages play an important role in xenogenic rejection and therefore may represent a major obstacle in clinical application of xenograft. CD33-related sialic acid–binding immunoglobulin–like lectins (Siglecs) belong to the immunoglobulin superfamily and contain a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM) that is able to inhibit cytokine production. Because human macrophages express various CD33-related Siglecs, we hypothesized that overexpression of α-2,6-sialyltransferase (2,6-ST) in swine endothelial cells (SECs) might prevent the cytotoxicity mediated by macrophages. To confirm our hypothesis, the cytotoxicity of macrophages against 2,6-ST–overexpressing SECs was determined with the use of in vitro–generated macrophages as an effector and naïve or 2,6-ST–overexpressing SECs as a target. The 2,6-ST–overexpressing SECs were established by transfection with the genes for 2,6-ST. Transfection of 2,6-ST led to significant reduction in cytotoxicity compared with naïve SECs. These findings indicate that the sialylated ligands against CD33-related Siglecs may provide an effective therapeutic strategy to inhibit macrophage-mediated xenograft rejection in xenotransplantation.     

Hepatectomy-Related Hypophosphatemia: A Novel Phosphaturic Factor in the Liver-Kidney Axis

Marked hypophosphatemia is common after major hepatic resection, but the pathophysiologic mechanism remains unknown. We used a partial hepatectomy (PH) rat model to investigate the molecular basis of hypophosphatemia. PH rats exhibited hypophosphatemia and hyperphosphaturia. In renal and intestinal brush-border membrane vesicles isolated from PH rats, Na⁺-dependent phosphate (Pi) uptake decreased by 50%–60%. PH rats also exhibited significantly decreased levels of renal and intestinal Na⁺-dependent Pi transporter proteins (NaPi-IIa [NaPi-4], NaPi-IIb, and NaPi-IIc). Parathyroid hormone was elevated at 6 hours after PH. Hyperphosphaturia persisted, however, even after thyroparathyroidectomy in PH rats. Moreover, DNA microarray data revealed elevated levels of nicotinamide phosphoribosyltransferase (Nampt) mRNA in the kidney after PH, and Nampt protein levels and total NAD concentration increased significantly in the proximal tubules. PH rats also exhibited markedly increased levels of the Nampt substrate, urinary nicotinamide (NAM), and NAM catabolites. In vitro analyses using opossum kidney cells revealed that NAM alone did not affect endogenous NaPi-4 levels. However, in cells overexpressing Nampt, the addition of NAM led to a marked decrease in cell surface expression of NaPi-4 that was blocked by treatment with FK866, a specific Nampt inhibitor. Furthermore, FK866-treated mice showed elevated renal Pi reabsorption and hypophosphaturia. These findings indicate that hepatectomy-induced hypophosphatemia is due to abnormal NAM metabolism, including Nampt activation in renal proximal tubular cells.Inorganic phosphate (Pi) absorption in the renal proximal tubules and small intestine is important for Pi homeostasis.¹ The Na⁺-dependent Pi (Na/Pi) transport system includes type IIa and type IIc Na/Pi transporters, which are localized in the apical membrane of the proximal tubular cells, and type IIb Na/Pi transporters, which are localized in the apical membrane of the intestinal epithelial cells.^1,2 Pi (re)absorption is regulated by the dietary Pi content, parathyroid hormone (PTH), and the active metabolite of vitamin D, 1α, 25-dihydroxyvitamin D3 [1,25(OH)₂D₃].³ Other phosphaturic hormones, termed phosphatonins, also control renal Pi handling.⁴ The discovery that fibroblast growth factor (FGF) 23, the first identified phosphatonin,⁵ originated from osteocytes established the concept of the bone-kidney axis.^6,7The incidence of liver transplantation has steadily increased and the incidence of partial hepatectomy (PH) has also consequently increased.⁸ Hypophosphatemia frequently occurs after liver resection.^9–11 Acute hypophosphatemia causes septicemia and is associated with a poor prognosis.^11,12 Acute hypophosphatemia is of considerable clinical relevance because many hepatectomized patients develop marked hypophosphatemia and, thus, large doses of Pi replacement are required to maintain metabolic homeostasis.¹³ Urinary Pi excretion is markedly increased in many patients. After hepatectomy, hypophosphatemia is associated with hyperphosphaturia.¹³For many years, the increased metabolic demand of the regenerating liver was considered the underlying pathologic mechanism of hypophosphatemia. The magnitude of Pi uptake by the recovering liver, however, cannot explain the severity of the resulting hypophosphatemia.¹¹ Hepatectomy-induced hypophosphatemia is associated with an increased renal fractional excretion index for Pi unrelated to intact FGF23, FGF7, or secreted frizzled-related protein 4 as a phosphaturic factor,¹⁴ indicating that other factors have a role in the pathogenesis of hypophosphatemia.Nicotinamide (NAM) inhibits intestinal and renal Na/Pi transport activity in normal rats.^15–17 Administration of NAM to rats produces a specific dose-dependent inhibition of Na/Pi transport across the renal brush-border membrane (BBM) and an increase in urinary Pi excretion.^16,17 NAM suppresses hyperphosphatemia in hemodialysis patients.¹⁸ Nicotinamide phosphoribosyltransferase (Nampt) catalyzes the first rate-limiting step in converting NAM to NAD,^19,20 which is essential for cellular metabolism, energy production, and DNA repair.^20–22 Nampt exists in two known forms: intracellular Nampt (iNampt) and secreted extracellular Nampt (eNampt).²³ eNampt also generates an intermediate product, nicotinamide mononucleotide (NMN).²³Our findings indicate that the acceleration of NAM metabolism through Nampt function in the kidney is involved in the hepatectomy-induced hypophosphatemia in rodent models. This study also suggests that NAM metabolism through the liver-kidney axis is important in Pi homeostasis.     

A retrospective comparative study of reconstructive methods following pancreaticoduodenectomy--pancreaticojejunostomy vs. pancreaticogastrostomy.

Reconstructive methods following pancreaticoduodenectomy in our department are discussed and evaluated in this study. Between January 1980 and November 1990 fifty-two consecutive patients underwent pancreaticoduodenectomy because of pancreas head disease. Thirty-one patients underwent pancreaticojejunostomy and twenty-one had pancreaticogastrostomy as reconstructive procedures. Mortality rate was 6% in pancreaticojejunstomy versus zero in pancreaticogastrostomy. Six patients had leakage from the pancreaticojejunostomy, but only one patient had necrosis of the gastric stump and leakage from the pancreaticogastrostomy. This case had previous distal gastrectomy done for gastric ulcer. The residual stomach might not have been large enough, and the blood supply of the gastric stump might not have been adequate for pancreaticogastrostomy. Except for this case, none was observed with leakage from the pancreatic anastomosis in the pancreaticogastrostomy group. No statistical significance in operating time or blood loss was observed between the two methods. The pancreaticogastrostomy cases without complications had significantly less loss of body weight than those with pancreaticojejunostomy at the date of discharge (p < 0.05). It is concluded that pancreaticogastrostomy is the safer reconstructive method following pancreaticoduodenectomy, although it may not be indicated in patients with prior gastrectomy.     

Transthyretin (prealbumin) in the pancreas and sera of newly diagnosed type I (insulin-dependent) diabetic patients.

We investigated transthyretin (TTR) in the pancreases and sera of 10 newly diagnosed type I diabetic patients by immunohistochemistry and nephelometry. In the type I diabetic pancreases, glucagon-positive A-cells showed strong immunoreactivity for TTR, the intensity and distribution pattern of which corresponded to those in normal subjects. Morphometric analysis revealed that the amount of strongly TTR-positive A-cells was not significantly different from that in normal subjects. On the contrary, insulin-positive B-cells, which normally show uneven and weak TTR immunoreactivity, decreased in number, and only a few residual B-cells showed faint immunoreactivity. Neither somatostatin cells nor pancreatic polypeptide cells were positive for TTR. The serum TTR concentration showed a significant decrease in type I diabetic patients compared with that in normal subjects (P less than 0.005). These data suggest that the synthesis or storage of TTR in A-cells is not affected, but that in B-cells is impaired in type I diabetes. The decrease in serum TTR might be one of the features of metabolic disorders in type I diabetes.