CD147 silencing inhibits lactate transport and reduces malignant potential of pancreatic cancer cells in in vivo and in vitro models.

BACKGROUND: CD147 (basigin, EMMPRIN) is a multifunctional, highly conserved glycoprotein enriched in pancreatic ductal adenocarcinomas (PDACs) which is associated with poor prognosis in many malignancies. The role of CD147 in pancreatic cancer, however, remains elusive. METHODS AND RESULTS: Silencing of CD147 by RNA interference (RNAi) reduced the proliferation rate of MiaPaCa2 and Panc1 cells. CD147 is required for the function and expression of the monocarboxylate transporters MCT1 and MCT4 that are expressed in human PDAC cells as demonstrated by real-time reverse transcription-PCR (RT-PCR) as well as immunohistology. MCT1 and MCT4 are the natural transporters of lactate, and MiaPaCa2 cells exhibited a high rate of lactate production, which is characteristic for the Warburg effect, an early hallmark of cancer that confers a significant growth advantage. Further induction of lactate production by sodium azide in MiaPaCa2 cells increased MCT1 as well as MCT4 expression. CD147 silencing inhibited the expression and function of MCT1 and MCT4 and resulted in an increased intracellular lactate concentration. Addition of exogenous lactate inhibited cancer cell growth in a dose-dependent fashion. In vivo, knock-down of CD147 in MiaPaCa2 cells by inducible short hairpin RNA (shRNA)-mediated CD147 silencing reduced invasiveness through the chorioallantoic membrane of chick embryos (CAM assay) and inhibited tumourigenicity in a xenograft model in nude mice. CONCLUSION: The function of CD147 as an ancillary protein that is required to sustain the expression and function of MCT1 and MCT4 is involved in the association of CD147 expression with the malignant potential of pancreatic cancer cells exhibiting the Warburg effect.

Fibrogenesis in the pancreas.

In recent years, numerous studies have provided novel insights into the pathomechanisms of pancreatic fibrogenesis. This includes in particular the identification and characterization of the pancreatic stellate cells (PSCs) and their role in the synthesis of extracellular matrix (ECM) proteins. It has become clear that pancreatic stellate cell activation is regulated by a complex network of growth factors and cytokines and results in increased expression and release of collagens I and II, fibronectin and other components of ECM. Among the cytokines involved in PSC activation and other fundamental mechanisms of pancreatic fibrosis, transforming growth factor beta (TGFbeta) is of particular relevance. TGFbeta stimulates PSC activation and induces transcription of ECM proteins mainly via activation of the Smad proteins which regulate gene expression through functional interaction with co-operating partner proteins such as the zinc finger transcription factor Sp1. Recent progress in understanding of the biochemical and molecular mechanisms of pancreatic fibrosis, is reviewed here.

Oxidized low-density lipoproteins bind to the scavenger receptor, CD36, of hepatic stellate cells and stimulate extracellular matrix synthesis.

Cumulating evidence suggests that oxidative stress resulting in lipid peroxidation and protein modification is involved in the pathogenesis of chronic liver injury and fibrogenesis. We investigated the effects of oxidized low-density lipoproteins (oxLDL) on collagen and fibronectin synthesis of cultured human and rat hepatic stellate cells (HSC). As shown on protein and mRNA levels, oxLDL dose-dependently stimulated the synthesis of collagen types I and III and fibronectin of cultured HSC. The effect was biphasic, with a maximum between 5 and 25 microg/mL oxLDL (c-fibronectin concentration in HSC supernatants increased 3.9-fold; collagen type I increased 4-fold). Higher oxLDL concentrations were cytotoxic. LDL modified with malondialdehyde (MDA) was not toxic, but stimulated extracellular matrix synthesis as well. As demonstrated by immunofluorescence microscopy (double staining of CD36 and iso-alpha-smooth muscle actin [iso-alpha-sm actin]), immunoblot, and reverse-transcription polymerase chain reaction (RT-PCR), respectively, cultured human HSC express the oxLDL receptor, CD36 (glycoprotein IIIb). Colocalization of CD36 and iso-alpha-sm actin on sinusoidal lining cells was further demonstrated using sections of human fibrotic liver. Preincubation of cultured human HSC with the monoclonal antibody, OKM5, known to block CD36-mediated oxLDL uptake, resulted in a reduction of the oxLDL-stimulated collagen type I synthesis by 56%. In summary, our results demonstrate that low concentrations of modified lipoproteins (oxLDL and MDA-LDL) represent fibrogenic mediators that bind to CD36 and stimulate matrix synthesis of HSC.

[Acute renal failure due to hypercalcemia and peripheral polyneuropathy--rare manifestations of sarcoidosis]. / Akutes Nierenversagen auf dem Boden einer Hyperkalzämie und periphere Polyneuropathie--seltene Manifestationen einer Sarkoidose.

BACKGROUND: Sarcoidosis can involve many organs. Whereas lung and intrathoracal lymph nodes are most often effected, there are some rare manifestations as renal failure and neuropathy. CASE REPORT: A 57-year-old male patient was referred to hospital with dyspnea on exertion, hypercalcemia and acute renal failure after holidays in Southern Europe. The diagnosis of sarcoidosis with the extraordinary manifestations of a nephropathia based on hypercalcemia and a sensitive polyneuropathy was made. CONCLUSION: These findings suggest that an asymptomatic sarcoidosis can develop hypercalcemia with renal failure by increased ultraviolet exposure. Patients with sarcoidosis should avoid marked exposure to UV light.

Depending on their concentration oxidized low density lipoproteins stimulate extracellular matrix synthesis or induce apoptosis in human coronary artery smooth muscle cells.

Various lines of evidence indicate that oxidative stress resulting in lipid peroxidation and protein modification is involved in the pathogenesis of atherosclerosis and coronary heart disease. We have investigated the effect of modified (oxidized) low-density lipoproteins (oxLDL) on collagen and fibronectin synthesis in cultured human coronary artery smooth muscle cells (HCA-SMC). As shown by immunofluorescence microscopy and time-resolved fluorescence immunoassay, oxLDL dose-dependently stimulated collagen type I and fibronectin synthesis in cultured HCA-SMC. The effect on matrix synthesis was biphasic, with a maximum effect at concentrations between 1 and 10 microg/ml oxLDL. Higher oxLDL concentrations (>25 microg/ml) were cytotoxic. Beside oxLDL, malondialdehyde-modified LDL also stimulated extracellular matrix synthesis. In the presence of 100 microg/ml ascorbic acid, 25, 50 and 100 microg/ml oxLDL induced apoptosis within 6-8 hours (demonstrated by TUNEL-reaction, annexin-V binding and APO-2.7-expression). Apoptosis was not induced by normal (unmodified) LDL and malondialdehyde-modified LDL. The radical scavengers and antioxidants TROLOX and probucol and the hydrogen peroxide eliminator catalase significantly reduced oxLDL-induced apoptosis. Our results demonstrate that low concentrations of oxLDL are profibrogenic by stimulating extracellular matrix synthesis, whereas higher oxLDL concentrations induce oxidative stress and apoptosis in coronary artery smooth muscle cells. The profibrogenic effect might be relevant in the formation of atherosclerotic plaques, and the proapoptotic effect might contribute to an increased plaque vulnerability.

Nephrotoxicity of aminophenols: effects of 4-dimethylaminophenol on isolated rat kidney tubules.

In isolated rat kidney tubules DMAP was found to inhibit the gluconeogenesis from lactate, pyruvate, or dihydroxyacetone. The ratio DMAP/protein rather than the calculated concentration of DMAP determined the strength of the effect, 20--25 nmoles DMAP/mg protein inhibiting the rate of gluconeogenesis by about 50%. The inhibition was not reversible. Phenacetin, 4-aminophenol and 4-acetamidophenol were much less effective than DMAP in inhibiting gluconeogenesis in isolated rat kidney tubules. DMAP 14C-labeled in the ring was quickly bound to proteins in kidney tubules. A portion of DMAP which did not exceed about 4 nmoles/mg protein, was bound in compounds soluble in perchloric acid. From this portion tris-GS-DMAP was isolated. DMAP diminished the glutathione content of isolated rat kidney tubules. Reduced glutathione added before DMAP prevented the inhibition of gluconeogenesis and diminished the binding of DMAP to proteins. The binding of DMAP required oxygen and was inhibited by carbon monoxide or cyanide. Several enzymes from isolated kidney tubules were found to be inhibited by DMAP doses which inhibited gluconeogenesis. Large DMAP doses also diminished the sums of ATP + ADP + AMP as well as NAD + NADH and NADP + NADPH. This effect corresponded to an increase in nucleotide degradation products and to increased activity of extracellular LDH. The results indicate that the inhibition of gluconeogenesis by DMAP is not due to a specific effect on one enzyme or on membranes but to unspecific reactions with many substances.