Role of extracellular matrix structural components and tissue mechanics in the development of postoperative pancreatic fistula

Radical resection remains the only curative treatment option in pancreatic cancer. Postoperative pancreatic fistulas (POPF) occur in up to 30 of patients leading to prolonged hospital-stay, increased cost of care and morbidity and mortality. Mechanical properties of the pancreas are associated with POPF. The aim of this study is to analyze the role of extracellular matrix (ECM) and tissue mechanics in the risk of POPF. Biopsies of 41 patients receiving a partial pancreas-resection are analyzed. Clinical data, ECM components and mechanical properties are correlated with POPF. Preoperative cholestasis is correlated with reduced risk of POPF, which comes along with a dilatation of the pancreatic duct and significantly higher content of collagen I. Patients developing POPF exhibited a degenerated tissue integrity, with significantly lower content of fibronectin and a trend for lower collagen I, III, IV and hyaluronic acid. This correlated with a soft tactile sensation of the surgeon during the intervention. However, this was not reflected with tissue mechanics evaluated by ex vivo uniaxial compression testing, where a significantly higher elastic modulus and no effect on the stress relaxation time were found. In conclusion, patients with cholestasis seem to have a lower risk for POPF, and an increase in collagen I. A degenerated matrix with lower content of structural ECM components correlates with increased risk of POPF. However, ex vivo uniaxial compression testing failed to clearly explain the link of ECM properties and POPF

Investigation of the Anti-Leishmania (Leishmania) infantum Activity of Some Natural Sesquiterpene Lactones

Leishmaniases are neglected infectious diseases caused by parasites of the ‘protozoan’ genus Leishmania. Depending on the parasite species, different clinical forms are known as cutaneous, muco-cutaneous, and the visceral leishmaniasis (VL). VL is particularly fatal and the therapy presents limitations. In the search for new anti-leishmanial hit compounds, seven natural sesquiterpene lactones were evaluated against promastigotes and intracellular amastigotes of Leishmania (Leishmania) infantum, a pathogen causing VL. The pseudoguaianolides mexicanin I and helenalin acetate demonstrated the highest selectivity and potency against intracellular amastigotes. In addition, promastigotes treated with helenalin acetate were subject to an ultrastructural and biochemical investigation. The lethal action of the compound was investigated by fluorescence-activated cell sorting and related techniques to detect alterations in reactive oxygen species (ROS) content, plasma membrane permeability, and mitochondrial membrane potential. Helenalin acetate significantly reduced the mitochondrial membrane potential and the mitochondrial structural damage was also confirmed by transmission electron microscopy, displaying an intense organelle swelling. No alteration of plasma membrane permeability or ROS content could be detected. Additionally, helenalin acetate significantly increased the production of nitric oxide in peritoneal macrophages, probably potentiating the activity against the intracellular amastigotes. Helenalin acetate could hence be a useful anti-leishmanial scaffold for further optimization studies

Paracrine interactions between mesenchymal stem cells affect substrate driven differentiation toward tendon and bone phenotypes

We investigated substrate dependent paracrine signaling between subpopulations of bone marrow stromal cells (BMSCs) that may affect the formation, or perhaps malformation, of the regenerating tendon to bone enthesis. Polyacrylamide substrates approximating the elastic modulus of tendon granulation tissue and the osteoid of healing bone (10-90 kPa) were functionalized with whole length fibronectin (Fn), type-I collagen (Col), or a mixed ligand solution (Fn/Col), and BMSCs were cultured in growth media alone or media supplemented with soluble Col or Fn. More rigid substrates with a narrow mechanical gradient (70-90 kPa) robustly induced osteogenic cell differentiation when functionalized with either Col or Fn. On broader mechanical gradient substrates (with a linear elastic modulus gradient from 10-90 kPa), cell differentiation was markedly osteogenic on subregions of Fn functionalized substrates above 20 kPa, but osteogenic activity was inhibited on all subregions of Col substrates. Osteogenic behavior was not observed when cells were cultured on Fn substrates if Col was present either in the media or on the substrate (Fn/Col). Tenogenic differentiation markers were observed only on Col substrates with moderate rigidity (∼30-50 kPa). Tenogenic differentiation was unaltered by soluble or substrate bound Fn. Co-culture of narrow gradient subsections revealed that any inclusion of tenogenic substrates (30-50 kPa, Col), caused otherwise osteogenic substrates to not develop markers of osteogenic differentiation, while increasing cell proliferation. These apparently paracrine effects could be mediated by bone morphogenetic protein-2 (BMP-2), as first confirmed by gene-level expression of BMP-2 and the transcription factor Smad8, and verified by BMP-2 media supplementation at levels similar to observed cell-secreted concentrations, which arrested osteogenic differentiation in 14 day cultures. Thus, cell instructive biomaterials with engineered mechanical and biochemical properties represent potentially powerful tools for directing BMSC differentiation to tendon and bone, however paracrine signals from tenogenic cells may delay osteogenesis at the healing enthesis