Mesenchyme-free expansion and transplantation of adult alveolar progenitor cells: steps toward cell-based regenerative therapies.

Alveolar type-2 (AT2) cells are necessary for the lung's regenerative response to epithelial insults such as influenza. However, current methods to expand these cells rely on mesenchymal co-culture, complicating the possibility of transplantation following acute injury. Here we developed several mesenchyme-free culture conditions that promote growth of murine AT2 organoids. Transplanting dissociated AT2 organoids into influenza-infected mice demonstrated that organoids engraft and either proliferate as AT2 cells or unexpectedly adopt a basal cell-like fate associated with maladaptive regeneration. Alternatively, transplanted primary AT2 cells also robustly engraft, maintaining their AT2 lineage while replenishing the alveolar type-1 (AT1) cell population in the epithelium. Importantly, pulse oximetry revealed significant increase in blood-oxygen saturation in primary AT2 recipients, indicating that transplanted cells also confer increased pulmonary function after influenza. We further demonstrated that both acid installation and bleomycin injury models are also amenable to AT2 transplantation. These studies provide additional methods to study AT2 progenitor potential, while serving as proof-of-principle for adoptive transfer of alveolar progenitors in potential therapeutic applications.

The effect of impaired angiogenesis on intestinal function following massive small bowel resection.

PURPOSE: Intestinal adaptation involves villus lengthening, crypt deepening, and increased capillary density following small bowel resection (SBR). Mice lacking the proangiogenic chemokine CXCL5 have normal structural adaptation but impaired angiogenesis. This work evaluates the impact of incomplete adaptive angiogenesis on the functional capacity of the intestine after SBR. METHODS: CXCL5 knockout (KO) and C57BL/6 wild-type (WT) mice underwent 50% SBR. Magnetic resonance imaging measured weekly body composition. Intestinal absorptive capacity was evaluated through fecal fat analysis. Gene expression profiles for select macronutrient transporters were measured via RT-PCR. Postoperative crypt and villus measurements were assessed for structural adaptation. Submucosal capillary density was measured through CD31 immunohistochemistry. RESULTS: Comparable postoperative weight gain occurred initially. Diminished weight gain, impaired fat absorption, and elevated steatorrhea occurred in KO mice after instituting high-fat diet. Greater postoperative upregulation of ABCA1 fat transporter occurred in WT mice, while PEPT1 protein transporter was significantly downregulated in KO mice. KO mice had impaired angiogenesis but intact structural adaptation. CONCLUSION: After SBR, KO mice display an inefficient intestinal absorption profile with perturbed macronutrient transporter expression, impaired fat absorption, and slower postoperative weight gain. In addition to longer villi and deeper crypts, an intact angiogenic response may be required to achieve functional adaptation to SBR.

Depletion of extracellular RANTES during human cytomegalovirus infection of endothelial cells.

Human cytomegalovirus (CMV) infection results in pneumonitis in bone-marrow and lung-transplant recipients. The source of CMV infection contributing to the onset of pneumonitis is unclear, but may involve infection of the lung endothelium in the presence of infiltrating mononuclear cells. Viral infection stimulates the host cell to express chemokines as signals to recruit specific immune cells to the site of injury. CMV encodes a chemokine receptor that may function to reduce host cell expression of chemokines. In the study reported here we found that extracellular concentrations of the chemokine regulated on activation, normal T cell expressed and secreted (RANTES) are depleted during productive infection of primary endothelial cells with CMV strain 4010, an endothelial-adapted strain of CMV. Utilizing adenovirus-transformed human kidney epithelial cells (type 293 cells) that stably express the CMV-encoded chemokine receptor US28, we found that depletion of extracellular RANTES during infection is attributable to US28, which binds and internalizes extracellular RANTES.