Side population cells derived from adult human liver generate hepatocyte-like cells in vitro.

We sought to determine whether hepatic side population (SP) cells derived from adult human liver possess the potential of a novel candidate hepatic stem cell. Human cadaveric donor liver was subjected to collagenase perfusion and hepatocytes were separated from nonparenchymal cells by differential centrifugation. SP cells were isolated from the nonparenchymal portion after Hoechst 33342 staining. Since CD45 is a panleukocyte antigen, CD45-negative SP cells were separated from the vast majority of CD45-positive SP cells (90%), and hepatic growth medium was used to culture both groups. Both CD45-negative and CD45-positive hepatic SP cells generated colonies in the hepatic growth medium in 2-3 weeks. The colonies yielded large cells morphologically consistent with human hepatocytes, demonstrating granule-rich cytoplasm, dense, often double nuclei, and intracellular lipofuscin pigment. The cultured cells from both sources were positive for markers of human hepatocytes: HepPar, cytokeratin 8 (CK8), and human albumin. Reverse transcriptase-polymerase chain reaction (RT-PCR) performed on both groups demonstrated positivity for additional liver markers including human albumin, CK18, alpha-1 anti-trypsin, and the human cytochrome P450 enzyme CYP2B6. Double immunostaining (CD45 and HepPar) and RT-PCR confirmed that the hepatocyte-like cells derived from the CD45-negative SP cells acquired HepPar positivity but had no detectable CD45 antigen expression. In contrast, the cultured cells derived from the CD45-positive SP cells also acquired HepPar positivity, but only a minimal fraction expressed the CD45 antigen. We conclude that hepatic SP cells derived from the nonparenchymal portion of human liver are a potential source of human hepatocytes irrespective of their CD45 status, and further animal studies will be required to assess their regenerative potential.

A review of achalasia in 33 children.

We reviewed our experience in the diagnosis and management of esophageal achalasia in 33 children over a 25-year period at a single center by a retrospective chart review of all patients diagnosed with achalasia between December 1, 1975 and January 30, 2001. There were 33 cases ranging from 5 months to 16 years of age at the time of presentation (17 boys and 16 girls). Although dysphagia and vomiting were the commonest presenting symptoms, weight loss, chest pain, coughing, and recurrent pneumonia also occurred in many patients. Barium contrast study of the esophagus was the initial diagnostic modality followed by esophageal manometry. An upper endoscopy was also performed in 78.7% of cases. Management was predominantly surgical; however, seven recently diagnosed patients opted for botulinum toxin (botox) injection as the first line of treatment. The follow-up duration varied from 10 months to 10 years (mean 4.71 +/- 3.2 years). Postsurgical complications included gastroesophageal reflux disease in five patients who had not received a simultaneous antireflux procedure and "residual achalasia" in two patients, who both responded to a single botox injection.