The rs738409 (I148M) variant of the PNPLA3 gene and cirrhosis: a meta-analysis

The human patatin-like phospholipase domain-containing-3 (PNPLA3) gene rs738409 C>G polymorphism is associated with several types of liver disease. The aim of this meta-analysis was to assess the risk of cirrhosis on the basis of rs738409 allele frequency and genotype. Medline, the Cochrane Library, EMBASE, and Google Scholar were searched for prospective and retrospective studies assessing the effect of the rs738409 polymorphism on liver cirrhosis. Seven studies, involving 2,023 patients with cirrhosis, were included. The G allele was associated with a significantly increased risk of cirrhosis versus the C allele [pooled odds ratio (OR) = 1.86, 95% confidence interval (CI) = 1.64–2.12, Z = 9.55, P < 0.001]. Both the GC and GG genotypes were associated with a significantly increased risk of cirrhosis versus the CC genotype (GC vs. CC: pooled OR = 1.73, 95% CI = 1.51–1.98, Z = 7.86, P < 0.001; GG vs. CC: pooled OR = 3.41, 95% CI = 2.77–4.18, Z = 11.65, P < 0.001). There was no evidence of publication bias. Our findings suggest that patients at risk for liver cirrhosis may benefit from PNPLA3 genotyping and thus more intensive monitoring if the rs738409 C>G polymorphism is identified.     

Association of PNPLA3 rs738409 polymorphism with liver steatosis but not with cirrhosis in patients with HBV infection: Systematic review with meta‐analysis

Background

Hepatitis B virus (HBV) infection is a worldwide health issue and is well known for being the main cause of developing secondary liver complications such as cirrhosis and hepatocellular carcinoma (HCC). The PNPLA3 rs738409 polymorphism has been investigated conclusively with occurrence risk of steatosis and cirrhosis. Therefore, performing a meta‐analysis of the available studies with the aim of clarifying the association between rs738409 and occurrence risk of steatosis and cirrhosis among HBV‐infected patients would be helpful.

Methods

Chronic HBV infection was defined as the persistence of HBsAg for more than 6 months. To gather sufficient data for this meta‐analysis, reliable databases were conclusively searched using appropriate keywords. Only studies that satisfied the inclusion criteria were enrolled in the present study.

Results

This meta‐analysis pooled four studies with 1135 cases of chronic hepatitis B (CHB) to evaluate the impact of PNPLA3 SNP on liver steatosis and also pooled five studies with 3713 cases of CHB to evaluate the impact of PNPLA3 SNP on cirrhosis. The association of rs738409 with each complication was investigated. The rs738409 was found to be associated with steatosis in recessive [p = 4.57 × 10^–6, odds ratio (OR) = 2.85], dominant (p = 4.35 × 10^‐6, OR = 1.84), co‐dominant (p = 6.18 × 10^‐8; OR = 3.74) and allelic (p = 9.79 × 10^‐9; OR = 1.78) models. No association was found between rs738409 and cirrhosis development in recessive (p = 0.99, OR = 1.00), dominant (p = 0.30, OR = 0.92), co‐dominant (p = 0.74; OR = 0.96) and allelic (p = 0.45; OR = 0.96) models.

Conclusions

Although the PNPLA3 rs738409 G allele has been associated with the risk of steatosis in CHB patients, no association between this polymorphism and the risk of cirrhosis was seen.     

The Impact of PNPLA3 rs738409 SNP on Liver Fibrosis Progression,Portal Hypertension and Hepatic Steatosis in HIV/HCV Coinfection

Background

Faster fibrosis progression and hepatic steatosis are hallmarks of HIV/HCV coinfection. A single nucleotide polymorphism (SNP) of the PNPLA3-gene is associated with development of non-alcoholic steatohepatitis and a worse outcome in alcoholic liver disease. However, the role of PNPLA3 rs738409 SNP on liver fibrosis and steatosis, portal hypertension, and virological response in HIV/HCV coinfection remains unclear.

Methods

In this cross-sectional study PNPLA3 (rs738409) and IL28B (rs12979860) SNPs were determined in 177 HIV/HCV coinfected patients. Liver fibrosis and steatosis—staged by liver biopsy and transient elastography using the Controlled Attenuation Parameter (CAP)–and portal hypertension (hepatic venous pressure gradient, HVPG) were compared across PNPLA3 genotypes.

Results

75 (42.4%) patients tested positive for a PNPLA3 minor/major risk allele (G/C:66; G/G:9) showed comparable fibrosis stages (median F2 vs. F2; p = 0.292) and similar amounts of hepatic steatosis (CAP: 203.5±41.9 vs. 215.5±59.7dB/m; p = 0.563) as compared to patients without a PNPLA3 risk allele. Advanced liver fibrosis was neither associated with PNPLA3 (p = 0.253) nor IL28B-genotype (p = 0.628), but with HCV-GT3 (p = 0.003), higher BMI (p = 0.008) and higher age (p = 0.007). Fibrosis progression rate (0.27±0.41 vs. 0.20±0.26 units/year; p = 0.984) and HVPG (3.9±2.6 vs. 4.4±3.0 mmHg; p = 0.472) were similar in patients with and without PNPLA3 risk alleles. SVR rates to PEGIFN/RBV therapy were similar across PNPLA3 genotypes.

Conclusions

The presence of a PNPLA3 risk allele had no independent impact on liver disease or virological response rates to PEGIFN/RBV therapy in our cohort of HIV/HCV coinfected patients.     

PARP1 rs1136410 C/C genotype associated with an increased risk of esophageal cancer in smokers

Molecular Biology Reports - DNA repair system plays a crucial role in maintaining genomic integrity and stability and in protecting against cancer. Poly(ADP-ribose) polymerase 1 (PARP1) functions...     

The association of PNPLA3 variants with liver enzymes in childhood obesity is driven by the interaction with abdominal fat

Background and Aims

A polymorphism in adiponutrin/patatin-like phospholipase-3 gene (PNPLA3), rs738409 C->G, encoding for the I148M variant, is the strongest genetic determinant of liver fat and ALT levels in adulthood and childhood obesity. Aims of this study were i) to analyse in a large group of obese children the role of the interaction of not-genetic factors such as BMI, waist circumference (W/Hr) and insulin resistance (HOMA-IR) in exposing the association between the I148M polymorphism and ALT levels and ii) to stratify the individual risk of these children to have liver injury on the basis of this gene-environment interaction.

Methods

1048 Italian obese children were investigated. Anthropometric, clinical and metabolic data were collected and the PNPLA3 I148M variant genotyped.

Results

Children carrying the 148M allele showed higher ALT and AST levels (p = 0.000006 and p = 0.0002, respectively). Relationships between BMI-SDS, HOMA-IR and W/Hr with ALT were analysed in function of the different PNPLA3 genotypes. Children 148M homozygous showed a stronger correlation between ALT and W/Hr than those carrying the other genotypes (p: 0.0045) and, therefore, 148M homozygotes with high extent of abdominal fat (W/Hr above 0.62) had the highest OR (4.9, 95% C. I. 3.2–7.8, p = 0.00001) to develop pathologic ALT.

Conclusions

We have i) showed for the first time that the magnitude of the association of PNPLA3 with liver enzymes is driven by the size of abdominal fat and ii) stratified the individual risk to develop liver damage on the basis of the interaction between the PNPLA3 genotype and abdominal fat.     

Association between the epidermal growth factor rs4444903 G/G genotype and advanced fibrosis at a young age in chronic hepatitis C

Background

The epidermal growth factor (EGF) rs4444903 A > G polymorphism has been associated with the development of liver cancer, which commonly complicates cirrhosis of viral origin; however, whether this polymorphism might be associated with fibrosis progression in chronic viral hepatitis is unknown. The present study was performed to assess the allelic and genotypic frequencies of the rs4444903 A > G polymorphism in patients with chronic hepatitis C virus HCV infection and to ascertain whether this polymorphism might be an independent predictor of the degree of fibrosis.

Methods

An RFLP-PCR technique was used to genotype 645 patients (211 with cirrhosis); 528 were referred for the diagnosis and treatment of chronic hepatitis C, and 117 were transplanted for HCV-related end stage liver disease. A group of 428 healthy subjects served as a control. All the subjects were of Caucasian ethnicity.

Results

The EGF rs4444903 A > G polymorphism genotype frequencies in HCV chronic infected patients were as follows: A/A = 227 (35.3%), A/G = 328 (50.9%), and G/G = 90 (14.8%). Genotype frequencies were found to differ between patients with an Ishak staging score ? 2 (A/A = 117, A/G = 157, G/G = 34) and patients with a score > 2 (A/A = 110, A/G = 171, G/G = 56, p = 0.038). A highly significant linear relationship between increasing stage scores and EGF genotype was detected in younger patients (A/A: 2.02 ± 0.18, A/G: 2.55 ± 0.17, G/G: 3.00 ± 0.32, p = 0.008). However, no significant association was detected between the stage score and EGF genotype in older patients (A/A: 3.79 ± 0.19, A/G: 3.64 ± 0.15, G/G: 3.98 ± 0.30 p = 0.579).

Conclusions

The EGF rs4444903 A > G polymorphism may facilitate liver fibrosis progression in Caucasian patients with chronic hepatitis C, especially in younger patients.     

Lamin A/C is a risk biomarker in colorectal cancer

Background

A-type lamins are type V intermediate filament proteins encoded by the gene LMNA. Mutations in LMNA give rise to diverse degenerative diseases related to premature ageing. A-type lamins also influence the activity of the Retinoblastoma protein (pRb) and oncogenes such a β-catenin. Consequently, it has been speculated that expression of A-type lamins may also influence tumour progression.

Methodology/Principal Findings

An archive of colorectal cancer (CRC) and normal colon tissue was screened for expression of A-type lamins. We used the Cox proportional hazard ratio (HR) method to investigate patient survival. Using CRC cell lines we investigated the effects of lamin A expression on other genes by RT-PCR; on cell growth by FACS analysis; and on invasiveness by cell migration assays and siRNA knockdown of targeted genes. We found that lamin A is expressed in colonic stem cells and that patients with A-type lamin-expressing tumours have significantly worse prognosis than patients with A-type lamin negative tumours (HR = 1.85, p = 0.005). To understand this finding, we established a model system based upon expression of GFP-lamin A in CRC cells. We found that expression of GFP-lamin A in these cells did not affect cell proliferation but did promote greatly increased cell motility and invasiveness. The reason for this increased invasiveness was that expression of lamin A promoted up-regulation of the actin bundling protein T-plastin, leading to down regulation of the cell adhesion molecule E-cadherin.

Conclusions

Expression of A-type lamins increases the risk of death from CRC because its presence gives rise to increased invasiveness and potentially a more stem cell-like phenotype. This report directly links A-type lamin expression to tumour progression and raises the profile of LMNA from one implicated in multiple but rare genetic conditions to a gene involved in one of the commonest diseases in the Western World.     

The association between interleukin-21 (rs2055979G/T) gene polymorphism and the risk of hepatocellular carcinoma and metastasis in patients with hepatitis C virus

Cancer prevalence is critically increasing worldwide; accordingly, improved prediction and therapeutic tools are necessary. Interleukin (IL)-21 is a potent antitumor cytokine, and the relationship between its gene variations and cancer risk is well established. Nevertheless, so far no study has investigated its role in hepatocellular carcinoma (HCC) progression and metastasis in hepatitis C virus (HCV)-infected people. Therefore, the present investigation was led on 267 Egyptian participants, involving 177 patients with HCV of which 90 patients had HCC (HCC group), 87 patients without HCC (non-HCC group), and 90 unrelated healthy controls. The association between rs2221903A/G and rs2055979G/T of the IL-21 gene and the risk of HCC and metastasis, as well as the clinico-pathological features, were analyzed. While rs2221903A/G polymorphism was not polymorphic in our cohort, patients carrying the genotype TT and allele T of the rs2055979G/T polymorphism had a significantly lower risk of HCC when comparing with HCC group and healthy controls. Also, participants carrying the aforementioned genotype and allele had a significantly lower risk of metastasis when comparing metastatic group with both nonmetastatic group and control group. The rs2055979G/T polymorphism was not significantly associated with clinico-pathological features of HCC. This is the first study to report a relationship between an intronic polymorphism in IL-21 gene and HCC and metastasis risk in the Egyptian people, in addition to identifying a potential new marker for the early detection and treatment of HCC.     

Prevalence of hepatitis C virus infection in Africa: Anti-HCV antibodies in the general population and in patients suffering from cirrhosis or primary liver cancer

Anti-hepatitis-C-virus (anti-HCV) antibody was tested for in sera from 410 adults living in Tunisia, Senegal, Burundi and Madagascar, and in 209 Tunisian and Senegalese patients suffering from liver diseases. Anti-HCV antibodies were detected in 4.2 % of the adult population from Africa, in 51 % of patients suffering from liver cirrhosis and in 37 % of patients suffering from primary liver cancer. However, higher proportions of anti-HCV antibodies were detected in HBsAg⁺ patients than in HBsAg⁻ patients. To assess the role of HCV in the development of both cirrhosis and primary liver cancer, a confirmation test is needed.     

The rat expresses two complement factor C4 proteins, but only one isotype is expressed in the liver

The complement component C4 is well known for its complex genetics in human and mouse where it is part of a tandemly duplicated module. For the rat, no such information had been available until recently. A C4 gene duplication could be identified also in the rat, but the duplicated module maps approximately 200 kb centromerically from the canonical C4-1 gene. In this study, we present the genomic organization of the two C4 gene-containing modules and the expression of the two C4 genes in the rat (Rattus norvegicus). The duplicated module contains an intact C4 gene as well as Cyp21 and Stk19 pseudogenes. Quantitative mRNA expression analyses revealed that both C4 genes are transcribed in various organs and tissues, but displaying ample differences of C4-1 and C4-2 expression. Most notably, C4-2 is not expressed in the liver. At variance to the mouse, the expression of the rat C4 genes does not exhibit any sex dependency. By using two-dimensional gel electrophoresis and mass spectrometry, products of both C4 genes could be identified in rat serum samples. These two rat C4 isotypes are nearly identical, but differ in a functionally important amino acid residue that is known to influence the functional properties of the C4 isotypes in human.     

MIF‐173G/C (rs755622) polymorphism as a risk factor for acute lymphoblastic leukemia development in children

Background

Macrophage inhibitory factor (MIF) is a pro‐inflammatory cytokine modulating monocyte motility and a pleiotropic regulator of different biological and cellular processes. The MIF‐173G/C (rs755622) polymorphism is found in the promoter region and affects its activity. The present study investigated the MIF polymorphism as a risk factor for the development of acute lymphoblastic leukemia (ALL) in Egyptian children.

Methods

We analyzed the MIF‐173G/C (rs755622) polymorphism in 180 ALL cases and 150 healthy control children by amplification of the gene using a polymerase chain reaction followed by restriction endonuclease digestion and running on an agarose gel for visualization of the product.

Results

We found a significant incidence of the homozygous polymorphic (CC) genotype and the combined polymorphic genotypes (GC + CC) in ALL patients compared to healthy controls (p = 0.001 and p = 0.007, respectively), whereas the wild‐type genotype (GG) was more common in healthy controls (p = 0.006). Multivariate logistic regression analysis adjustment for MIF different genotypes and other potential risk factors such as age, sex and parental smoking indicated that the CC genotype is the only significant risk factor for the test (p = 0.02). We also noted that, by increasing the C‐allele representation within the gene [GC, CC], there was an increase in total leukocytic count (p = 0.09 and p = 0.001, respectively) that may reflect the bad prognostic impact of the polymorphic allele, although further studies are needed.

Conclusions

The results of the present study indicate that the MIF‐173G/C (rs755622) polymorphism is a risk factor for childhood ALL development with respect to both homozygous and combined polymorphic genotypes. In addition, the increased leukocytic count in synchronization with the increased representation of the polymorphic C‐allele may reflect its bad prognostic impact.     

Radiation-induced genetic instability: no association with changes in radiosensitivity or cell cycle checkpoints in C3H 10T1/2 mouse fibroblasts

We investigated various phenotypic characteristics of radiation-induced morphologically transformed C3H 10T1/2 mouse fibroblasts. The cells were treated with 8 Gy x-rays, and type II/III foci were isolated. Cell lines were developed from these foci, and subsequently clones were established from these focal lines. The clones were examined for DNA content, radiosensitivity and inducible cell cycle arrests. Besides the morphological changes associated with the transformed state, the major difference between the isolated focal lines or derived clones and the parental C3H 10T1/2 line was one of ploidy. The transformed cells often displayed aneuploid and multiple polyploid populations. No change in the radiosensitivity of the transformed cells was observed. Furthermore, the two major radiation- and staurosporine-induced G1 and G2 cell cycle arrests observed in the parental cell line were also observed in the morphological transformants, suggesting that checkpoint function was normal. Received: 15 May 1997 / Accepted in revised form: 21 October 1997     

Microbe-specific C3b deposition in the horseshoe crab complement system in a C2/factor B-dependent or -independent manner

Complement C3 plays an essential role in the opsonization of pathogens in the mammalian complement system, whereas the molecular mechanism underlying C3 activation in invertebrates remains unknown. To understand the molecular mechanism of C3b deposition on microbes, we characterized two types of C2/factor B homologs (designated TtC2/Bf-1 and TtC2/Bf-2) identified from the horseshoe crab Tachypleus tridentatus. Although the domain architectures of TtC2/Bf-1 and TtC2/Bf-2 were identical to those of mammalian homologs, they contained five-repeated and seven-repeated complement control protein domains at their N-terminal regions, respectively. TtC2/Bf-1 and TtC2/Bf-2 were synthesized and glycosylated in hemocytes and secreted to hemolymph plasma, which existed in a complex with C3 (TtC3), and their activation by microbes was absolutely Mg(2+)-dependent. Flow cytometric analysis revealed that TtC3b deposition was Mg(2+)-dependent on Gram-positive bacteria or fungi, but not on Gram-negative bacteria. Moreover, this analysis demonstrated that Ca(2+)-dependent lectins (C-reactive protein-1 and tachylectin-5A) were required for TtC3b deposition on Gram-positive bacteria, and that a Ca(2+)-independent lectin (Tachypleus plasma lectin-1) was definitely indispensable for TtC3b deposition on fungi. In contrast, a horseshoe crab lipopolysaccharide-sensitive protease factor C was necessary and sufficient to deposit TtC3b on Gram-negative bacteria. We conclude that plasma lectins and factor C play key roles in microbe-specific TtC3b deposition in a C2/factor B-dependent or -independent manner.     

The CDKN2a common variants: 148 Ala/Thr and 500 C/G in 3' UTR, and their association with clinical course of melanoma

Changes in CDKN2a gene are known to be linked with sporadic melanoma and hereditary predisposition to this cancer. In the Polish population mutations in the coding region of the CDKN2a gene are rather rare, therefore the attention has been focused on polymorphisms and alterations in uncoding regions such as 3' UTR. The aim of this study was to analyze two common polymorphisms, Ala148Thr and 500 C/G, and correlate them with the clinical course of melanoma. DNA from 285 patients was analyzed and found polymorphisms were correlated with the clinical parameters employing statistical methods. The obtained results allow us to conclude: (i) survival times of 500 C/G carriers vs. cumulating proportion surviving was not statistically significant; (ii) CDKN2a polymorphism 500 C/G correlated with Ala148Thr; (iii) no correlation was observed between the 500 C/G polymorphism and age of diagnosis, localization of primary melanoma and survival time; (iv) we did not find correlation between 500 C/G and type of cancer in the family; (v) changes in the CDKN2a gene were not found in patients with second cancer.     

The transcription factor C/EBPbeta is essential for inducible expression of the cox-2 gene in macrophages but not in fibroblasts

Cyclooxygenase-2 (COX-2) is the rate-limiting enzyme for the inducible synthesis of prostaglandins, and its up-regulated activity is thought to play a pathological role in diseases such as inflammatory bowel disease, rheumatoid arthritis, and cancer. Regulation of COX-2 expression is complex and appears to involve diversified mechanisms in different cell types and conditions. Here we make use of immortalized macrophages and fibroblasts that we have generated from C/EBPbeta-deficient mice to directly test and compare the specific role played by this factor in inducible COX-2 expression in these two cell types. We could demonstrate that COX-2 mRNA induction and promoter activity were profoundly impaired in C/EBPbeta(-/-) macrophages and could be rescued by expression of C/EBPbeta. The obligatory role of C/EBPbeta in COX-2 expression appeared to be mediated exclusively by the C/EBP element located at positions -138/-130 of the murine cox-2 promoter, and did not involve altered activity at the level of the other promoter elements described previously (the -402/-392 NF-kappaB site, the -59/-48 CRE/E box element, and a potential second C/EBP site located at positions -93/-85). In contrast, COX-2 induction was completely normal in C/EBPbeta-deficient fibroblasts, thus highlighting the diversity of cell-specific molecular mechanisms in determining inducible COX-2 expression and prostaglandins production.     

EGF-induced MMP-9 expression is mediated by the JAK3/ERK pathway,but not by the JAK3/STAT-3 pathway in a SKBR3 breast cancer cell line

The number of epidermal growth factor receptors (EGFRs) and their ligands are highly expressed in malignant tumor cells. The EGF signaling pathway is also activated in up to one-third of patients with breast cancer. In this study, we investigated the novel function of the JAK3 inhibitor, WHI-P131, on EGF-induced MMP-9 expression and the regulatory mechanism of EGF-induced MMP-9 expression in SKBR3 cells. We observed that EGF increased MMP-9 mRNA and protein expression in a dose-dependent manner. EGF also induced the phosphorylation of EGFR, ERK, and STAT-3, and these effects were inhibited by the EGFR inhibitor, AG1478. To investigate the involvement of the STAT-3 pathway on EGF-induced MMP-9 expression, we pretreated SKBR3 cells with JAK1, JAK2, and JAK3 inhibitors prior to EGF treatment. The results showed that the JAK3 inhibitor, WHI-P131, as well as JAK3 siRNA transfection, but not the JAK1 and JAK2 inhibitors, significantly decreased EGF-induced MMP-9 expression. In addition, EGF-induced STAT-3 phosphorylation was only inhibited by WHI-P131. We then transfected cells with adenoviral STAT-3 (Ad-STAT-3), followed by treatment with EGF. Interestingly, EGF-induced MMP-9 expression was decreased by Ad-STAT-3 overexpression in a dose-dependent manner, while it was significantly increased by STAT-3 siRNA transfection. Our results also showed that basal levels of MMP-9 expression were significantly increased by constitutive active-MEK (CA-MEK) overexpression. EGF-induced ERK phosphorylation was prevented by WHI-P131, but not by JAK1 and JAK2 inhibitors. On the other hand, EGF-induced MMP-9 expression was decreased by the MEK1/2 inhibitor, UO126. Therefore, for the first time, we suggest that the JAK3 inhibitor, WHI-P131, inhibits EGF-induced STAT-3 phosphorylation as well as ERK phosphorylation. The JAK3/ERK pathway may play an important role in EGF-induced MMP-9 expression in SKBR3 cells.     

The association of MDR1 C3435T and G2677T/A polymorphisms with plasma platelet-activating factor levels and coronary artery disease risk in Turkish population

Increased levels of peripheral proinflammatory mediators can contribute to the development of coronary artery disease (CAD). Platelet activating factor (PAF) is an important proinflammatory mediator and plasma levels of PAF correlate with transmembrane transporter multidrug resistant 1 P-glycoprotein (MDR1 Pgp) expression and activity. MDR1 polymorphisms can affect the expression and activity of Pgp and plasma PAF levels. Therefore, we investigated the possible relationship between MDR1 C3435T and G2677T/A polymorphisms and plasma PAF levels and the risk of CAD. The study population consisted of 198 patients angiographically documented CAD, including 113 cases with at least 1 coronary artery with ≥ 50% luminal diameter stenosis and 85 control subjects with strictly normal coronary angiograms. Genotypes of the MDR1 C3435T and G2677T/A polymorphisms were determined by polymerase chain reaction (PCR) followed by restriction fragment length polymorphism (RFLP). Plasma PAF levels were detected by enzyme-linked immunosorbent assay (ELISA). There were no significant differences among plasma PAF levels in regard to MDR1 C3435T and G2677T polymorphisms in CAD patients and controls. No statistically significant difference was found for the genotypic and allelic distributions of the polymorphisms in the MDR1 gene between the patients and the control subjects. Furthermore, analysis of MDR1 haplotypes did not show any associations with increased plasma PAF levels and risk of CAD. Our results suggest that plasma PAF levels are not associated with MDR1 gene polymorphisms. There is no association between MDR1 C3435T and G2677T/A polymorphisms and the risk of CAD in Turkish patients.     

Stretch-induced retinal vascular endothelial growth factor expression is mediated by phosphatidylinositol 3-kinase and protein kinase C (PKC)-zeta but not by stretch-induced ERK1/2, Akt, Ras, or classical/novel PKC pathways.

Stretch-induced expression of vascular endothelial growth factor (VEGF) is thought to be important in mediating the exacerbation of diabetic retinopathy by systemic hypertension. However, the mechanisms underlying stretch-induced VEGF expression are not fully understood. We present novel findings demonstrating that stretch-induced VEGF expression in retinal capillary pericytes is mediated by phosphatidylinositol (PI) 3-kinase and protein kinase C (PKC)-zeta but is not mediated by ERK1/2, classical/novel isoforms of PKC, Akt, or Ras despite their activation by stretch. Cardiac profile cyclic stretch at 60 cpm increased VEGF mRNA expression in a time- and magnitude-dependent manner without altering mRNA stability. Stretch increased ERK1/2 phosphorylation, PI 3-kinase activity, Akt phosphorylation, and PKC-zeta activity. Signaling pathways were explored using inhibitors of PKC, MEK1/2, and PI 3-kinase; adenovirus-mediated overexpression of ERK, PKC-alpha, PKC-delta, PKC-zeta, and Akt; and dominant negative (DN) mutants of ERK, PKC-zeta, Ras, PI 3-kinase and Akt. Although stretch activated ERK1/2 through a Ras- and PKC classical/novel isoform-dependent pathway, these pathways were not responsible for stretch-induced VEGF expression. Overexpression of DN ERK and Ras had no effect on VEGF expression in these cells. In contrast, DN PI 3-kinase as well as pharmacologic inhibitors of PI 3-kinase blocked stretch-induced VEGF expression. Although stretch-induced PI 3-kinase activation increased both Akt phosphorylation and activity of PKC-zeta, VEGF expression was dependent on PKC-zeta but not Akt. In addition, PKC-zeta did not mediate stretch-induced ERK1/2 activation. These results suggest that stretch-induced expression of VEGF involves a novel mechanism dependent upon PI 3-kinase-mediated activation of PKC-zeta that is independent of stretch-induced activation of ERK1/2, classical/novel PKC isoforms, Ras, or Akt. This mechanism may play a role in the well documented association of concomitant hypertension with clinical exacerbation of neovascularization and vascular permeability.