Heparanase accelerates the proliferation of both hepatocytes and endothelial cells early after partial hepatectomy.

BACKGROUND AND AIMS: Heparanase (HPSE) is an endo-ß-D-glucuronidase, which cleaves heparan sulfate in the extracellular matrix (ECM) and has pro-angiogenic and pro-proliferative properties. The aim of this investigation was to study the effect of HPSE on hepatocytes and endothelial cells (EC) during liver regeneration. METHODS: Following 70% hepatectomy (PHP), rats were injected daily with 1-50µg HPSE/rat. Liver samples were stained with H&E and anti-bromodeoxyuridine (BrdU) antibody. mRNAs of hepatocyte growth factor (HGF), stem cell factor, tumor necrosis factor (TNF)-α, interleukin(IL)-6, and cyclinD1 were tested by real-time qPCR. Matrix metalloproteinases (MMPs) were tested by gel zymography. RESULTS: Compared to the saline control, HPSE increased hepatocyte proliferation 24h, 48h and 72h after PHP, with the maximal effect found at 24h with 50µg HPSE (40.9±2.5% vs. 8.6±4.3%, p<0.01 for BrdU staining; 5.5±0.9% vs. 0.8±0.5%, p<0.05 for mitosis). Proliferation of the sinusoidal and the portal vein radical ECs was also increased (p<0.05). HPSE caused a twofold increase in cyclinD1 mRNA (p<0.05) and in pro-MMP-9 levels (p<0.05). HPSE at all doses also caused significant reductions of TNF-α mRNA (p<0.05) and IL-6 mRNA, and no change in HGF mRNA. CONCLUSIONS: HPSE enhances liver regeneration by inducing proliferation of hepatocytes and both sinusoidal and vascular ECs. Since the effect of HPSE on hepatocytes occurred earlier than that observed in ECs, this effect is not related to HPSE's effect on ECs. The mechanism of HPSE action is probably indirect and is mediated by HPSE-dependent ECM cleavage and the release of pre-existing enzymes.

Near-missed upper tracheoesophageal fistula in esophageal atresia.

Upper pouch tracheoesophageal fistula (TEF) accompanying esophageal atresia (EA) occurs in less than 1% of all EA/TEF variants and could be easily missed after birth. To confront such diagnostic inaccuracy, perioperative tracheobronchoscopy (TBS) and preoperative upper pouch esophagogram (UPEG) have been proposed but are still controversial. We describe the role of UPEG and TBS, used early after birth, in two cases of EA/TEF with upper pouch TE fistulas with unusual high location (one intrathoracic, one subglotic). These upper TE fistulas were almost missed but ultimately detected very early while employing both UPEG and TBS, wherein UPEG was for the diagnosis of TEF and TBS for both intraoperative diagnostic confirmation and aid in TEF identification. We conclude that UPEG and TBS are complementary in detecting near-missed upper TE fistula accompanying EA. Such approach ensures early and accurate diagnosis of EA/TEF variants, thus preventing the complications of a missed congenital upper pouch TE fistula.

Heparanase enhances early hepatocyte inclusion in the recipient liver after transplantation in partially hepatectomized rats.

Hepatocyte transplantation is an emerging approach for the treatment of liver diseases. However, broad clinical application of this method has been limited by restricted source of cells and low efficiency of cell integration within the recipient liver. Heparanase cleaves heparan sulfate proteoglycans in the extracellular matrix and basement membrane, activity that affects cellular invasion associated with cancer metastasis and inflammation. This activity has a multifunctional effect on cell-cell interaction, cell adhesion, and angiogenesis. All these factors are important for successful integration of transplanted hepatocytes. Male donor hepatocytes pretreated with heparanase or untreated were transplanted into recipient female rat spleen following partial hepatectomy. Engraftment efficacy was evaluated by PCR for Y chromosome, histology and PCNA, and heparanase immunohistochemistry. In addition, proliferative activity of hepatocytes in vitro was determined by bromodeoxyuridine immunostaining. The number of heparanase-treated cells detected in the recipient liver was significantly increased three- to fivefold within 24-48 h posttransplantation and twofold at 14 days compared with untreated cells. The transplanted hepatocytes treated with heparanase were clearly seen inside portal vein radicles as cell aggregates up to 72 h posttransplantation. The number of portal radicles filled with heparanase-treated hepatocytes was increased compared to control early after transplantation. Heparanase treatment enhanced hepatocyte and sinusoidal endothelial cell proliferation in the liver, and hepatocyte proliferation within the spleen tissue. Preliminary in vitro studies with isolated hepatocytes treated with heparanase showed increased proliferative activity within 24-48 h of cell culture. These results suggest that preincubation of hepatocytes with heparanase increases the presence of hepatocytes within the recipient liver early following cell transplantation and stimulates both hepatocyte and sinusoidal endothelial cell proliferation.

Rectal prolapse following posterior sagittal anorectoplasty for anorectal malformations.

PURPOSE: Rectal prolapse is a known postoperative problem in children with anorectal malformations. The aims of this study were to determine the incidence of significant rectal prolapse (>5 mm), to objectively quantify its predisposing factors, and to offer recommendations as to its prevention and surgical treatment. METHODS: The authors reviewed their series of 1619 patients with anorectal malformations; 1169 underwent primary posterior sagittal anorectoplasty (PSARP) at their institution between 1980 and 2002, and complete records were available for 833. The series was analyzed for incidence of prolapse, type of anorectal malformation, status of the sacrum, muscle quality, associated vertebral and spinal anomalies, and postoperative constipation. A specific technique for prolapse repair was used. RESULTS: Of 833 patients, 45 developed significant rectal prolapse (3.8%). The mean age at the time of PSARP was 0.73 years (range, 0.19-5 years). The average time to recognition of prolapse following PSARP was 13.1 months. Of these 45 patients, 32 required surgical repair and of those, 3 required a second surgical repair. The incidence of prolapse varied by complexity of anorectal defect: cloaca (6.2%), rectobladder neck fistula (6.8%), rectourethral fistula (5.4%), rectovestibular fistula (1.2%), rectal atresia (0%), and rectoperineal fistula (0%). There was a significantly increased incidence of prolapse in patients with a low muscle quality score and in patients with vertebral anomalies (20% vs 3.2%). The presence of a tethered cord and an abnormal sacral ratio did not correlate with an increased incidence of prolapse. Twenty-two patients developed prolapse following colostomy closure, and of these, 12 (55%) suffered from constipation. CONCLUSIONS: The overall incidence of significant rectal prolapse following PSARP is low. Prevention of prolapse with the PSARP technique may be because of key technical steps. Patients with higher anorectal malformations, poorer muscle quality, and vertebral anomalies had a greater risk of developing postoperative rectal prolapse. The presence of tethered cord and quality of the sacrum were not predictive of postoperative prolapse. Constipation seems to be a factor in the development of prolapse.

Enhancing the vascularization of three-dimensional porous alginate scaffolds by incorporating controlled release basic fibroblast growth factor microspheres.

Site-specific delivery of angiogenic growth factors from tissue-engineered devices should provide an efficient means of stimulating localized vessel recruitment to the cell transplants and would ensure cell survival and function. In the present article, we describe the construction of a novel porous alginate scaffold that incorporates tiny poly (lactic-co-glycolic acid) microspheres capable of controlling the release of angiogenic factors, such as basic fibroblast growth factor (bFGF). The microspheres are an integral part of the solid alginate matrix, and their incorporation does not affect the scaffold porosity or pore size. In vitro, bFGF was released from the porous composite scaffolds in a controlled manner and it was biologically active as assessed by its ability to induce the proliferation of cardiac fibroblasts. The controlled delivery of bFGF from the three-dimensional scaffolds accelerated the matrix vascularization after implantation on the mesenteric membrane in rat peritoneum. The number of penetrating capillaries into the bFGF-releasing scaffolds was nearly fourfold higher than into the control scaffolds (those incorporating microspheric BSA and heparin but not bFGF). At day 10 posttransplantation, capillary density in the composite scaffolds was 45 +/- 3/mm(2) and it increased to 70 +/- 7/mm(2) by day 21. The released bFGF induced the formation of large and matured blood vessels, as judged by the massive layer of mural cells surrounding the endothelial cells. The control over bFGF delivery and localizing its effects to areas of need, may aid in the wider application of bFGF in therapeutic angiogenesis as well as in tissue engineering.