Abnormal crosstalk between pancreatic acini and macrophages during the clearance of apoptotic cells in chronic pancreatitis

Abstract

In chronic pancreatitis (CP), both the progressive loss of acinar parenchyma and aggressive fibro-inflammatory reactions ultimately lead to irreversible organ destruction. Dying cells are normally removed by macrophages and elimination is associated with anti-inflammatory cytokine switch. We investigated whether defective clearance of damaged acini by macrophages such as compromised phagocytosis or altered cytokine reaction occurs in CP and thus represents a causative link between acinar loss and fibro-inflammation. In a checkerboard-like co-culture system, we assessed normal and CP macrophages for their phagocytic and cytokine responses to dying pancreatic acinar cells of normal or CP origin by FACS, confocal microscopy, QRT-PCR, and ELISA. In CP, phagocytosis of apoptotic acini by macrophages was not impaired; however, the associated cytokine responses were gradually perturbed. Most interestingly, only normal acini suppressed TGFbeta1 expression and accumulation specifically in normal macrophage cultures, while CP acini lost this ability. Both types of apoptotic acini induced pro-inflammatory cytokine bursts of varying strength in both types of macrophages; however, the most significant difference (more than 50-fold higher expression of IL-1beta, IL-6, and IL-8) was evident between CP/CP and normal/normal combinations, indicating that acinar and macrophage alterations synergistically lead to the ultimate CP-specific bias. In combination with in situ data comparing circulating inflammatory cells to pancreatic resident ones, our results indicate that cytokine expression in inflammatory cells undergoes spatiotemporal modulation, most likely through a successive interplay of acinar, stromal, and circulating factors. Thus, clearance of injured pancreatic acini by macrophages is associated with a unique cytokine reaction which may constitute a basis for progression of SAPE (sentinel acute pancreatitis event) to the irreversible fibro-inflammation in CP.

Chondroitin Sulfate Proteoglycan CSPG4 as a Novel Hypoxia-Sensitive Marker in Pancreatic Tumors

“…All reagents and equipment for mRNA/cDNA preparation were supplied by Roche Applied Science (RAS, Mannheim, Germany) [23], [24], [26]. mRNA was prepared by automated isolation using a MagNA Pure LC instrument and isolation kit.…”

Immune cells participate in the oncosuppressive activity of parvovirus H-1PV and are activated as a result of their abortive infection with this agent

“…mRNA was prepared by automated isolation using the MagNA pure LC instrument and isolation kit I. cDNA was prepared using the first-strand cDNA synthesis kit for RT-PCR according to the manufacturer's instructions. 31 For RT-PCR total RNA was extracted from pancreatic tumors or celiac lymph nodes of treated animals and reverse transcribed into cDNA, as previously described. 32 The primers used to amplify rat IFNγ cDNAs were 5'-ATC TGG AGG AAC TGG CAA AAG GAC G-3'-forward and 5'-CCT TAG GCT AGA TTC TGG TGA CAG C-3'-reverse, resulting in a product of 290 bp.…”

Prevailing Role of Contact Guidance in Intrastromal T-cell Trapping in Human Pancreatic Cancer

“…Chemokine and CR mRNA levels in total tissue lysates were determined by quantitative real-time PCR (qRT-PCR) as described previously (28,29). All materials and equipment for RNA and cDNA preparation were obtained from Roche and used following the manufacturer's instructions.…”