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Abstract Increased GP73 expression in hepatocytes from patients with acute hepatitis, through disease progression to cirrhosis and chronic liver disease suggests that progressive tissue remodeling and fibrogenesis are driving forces for GP73 upregulation. Nevertheless, details about regulation of GP73 expression and its biological functions remain elusive and await further characterization. In this study, we demonstrate that GP73 is a direct target of TGF-β1 transcriptional regulation. Its induced expression inhibits TGF-β-Smad mediated growth suppression. On the other hand, elevated GP73 results in upregulation of ERK/Akt signaling induced by TGF-β1. Mechanistically, upregulation of lipid raft and caveolae-1 induced by GP73 overexpression mediates its regulatory effect on TGF-β1 signaling. Notably, lipid raft expression is elevated in HCC tumors and tissues with higher GP73 expression yield more intensive Flotillin staining. Our results establish the linkage between GP73 and TGF-β signaling, indicating that GP73 may promote HCC tumorigenesis by selectively regulating TGF-β signaling through lipid raft modulation. Highlights • GP73 expression induced by TGF-β1 transcriptional regulation inhibits TGF-β-Smad mediated growth suppression. • Elevated GP73 results in upregulation of extracellular signal-regulated kinases (ERK) signaling. • Lipid raft and caveolae upregulation induced by GP73 overexpression mediates its regulatory effect on TGF-β signaling. • Lipid raft expression is elevated in HCC tumors, tissues with higher GP73 expression yield more intensive Flotillin staining. • GP73 may selectively regulate TGF-β signaling via lipid raft modulation and probably promotes HCC tumorigenesis. [ABSTRACT FROM AUTHOR]

To better understand the molecular mechanism underlying of diapause in Antheraea pernyi (A.pernyi), we cloned a novel diapause-associated protein 3 (DAP3) gene from A.pernyi by reverse transcription-polymerase chain reaction (RT-PCR) and studied the biological functions. Sequence analysis revealed that this gene encodes 171 amino acids and has a conserved domain of Copper/Zinc superoxide dismutase (Cu/Zn-SOD). Western blot and qRT-PCR results showed that DAP3 was mainly expressed in the pupal stage, and gradually decreased as diapause development. DAP3 was also expressed in 1st and 5th instar larvae of A.pernyi. In tissues of 5th instar larvae of A.pernyi, DAP3 was mainly expressed in the epidermis, followed by the head, hemolymph and fat body. To identify the SOD activity of DAP3, we constructed a prokaryotic expression vector by inserting the coding region sequence into plasmid pET-28a (+) and obtained the purified rHIS-DAP3 fusion protein by Ni-NTA affinitive column. Importantly, we found the SOD activity of DAP3 fusion protein was approximately 0.6674 U/µg. To further confirm the SOD activity of DAP3 in vivo, we induced the oxidative stress model of pupae by UV irradiation. The results showed that both the mRNA and protein level of DAP3 significantly increased by UV irradiation. Furthermore, the SOD activity of the total lysate of pupae increased significantly at 10 min post UV irradiation and transiently returned to normal level afterwards. These results suggested that DAP3 might be a novel protein with SOD activity getting involved in regulation of diapause in A.pernyi. [ABSTRACT FROM AUTHOR]

Context.—Kallikreins are a subgroup of serine proteases with diverse physiologic functions. It has been confirmed that kallikrein 7 (KLK7) is differentially expressed in ovarian and breast cancer. Antileukoprotease (ALP) has been shown to be a specific inhibitor of human kallikrein 7 (hK7). Antileukoprotease overexpression is commonly associated with aggressive, high-risk, or metastatic cancer originating from various organs. Objective.—To investigate the expression and potential role of hK7 and its inhibitor ALP in prostate cancer. Design—The mRNA expression of KLK7 and ALP transcript in benign prostate epithelial cells and prostate cancers was evaluated by semiquantitative reverse transcription-polymerase chain reaction. We examined hK7 and ALP protein expression by immunohistochemistry in 20 normal prostate tissues, 50 benign prostatic hyperplasia tissues, and 103 prostate cancers. Western blot examination showed protein expression of hK7 and ALP in benign prostate epithelial cells and prostate cancer cell lines. Results.—Semiquantitative polymerase chain reaction examination revealed that the mRNA level of KLK7 and ALP was significantly decreased in prostate cancers compared with that in benign prostate epithelial cells (P < .001). Immunohistochemical expression of hK7 was observed in prostate epithelial cells, whereas little or no staining was observed in prostate cancer. Western blot analysis revealed that hK7 and ALP were decreased in malignant prostate epithelium. Conclusions—Like hK7, ALP is down-regulated in prostate cancers, which begs the question of whether it remains an effective inhibitor of hK7 or whether it is discordant in time or space and is ineffective as an inhibitor of hK7. The function of KLK7 and ALP in prostate cancer should be further studied. (Arch Pathol Lab Meó. 2OO8;132:1796-1S01 ) [ABSTRACT FROM AUTHOR]

Background. Excessive activation of the nod-like receptor family pyrin domain containing 3(NLRP3) inflammasome plays a significant role in the progression of cardiac injury. In China, it has been well recognized that Chinese herbal medicine is markedly effective in treating cardiovascular diseases (CVDs). LuQi Formula (LQF) has been used clinically for more than 10 years and confirmed to be effective in improving cardiac function and inhibiting apoptosis. However, the specific mechanisms underlying its efficacy are mostly unknown. This study aimed to evaluate whether LQF could alleviate cardiac injury and apoptosis by regulating the NLRP3 inflammasome and the caspase-3/Bax pathway. Purpose. In this study, we investigated the effects of LQF on cardiac remodeling in a mouse model of myocardial infarction (MI) in vivo. Methods. Forty male C57BL/6 mice were randomly divided into four groups: the sham group, the model group, the LQF group, and the perindopril group, with a sample size (n) of 10 mice in each group. Except the sham group, the other groups received left anterior descending (LAD) coronary artery ligation to induce MI and then treated with LQF, perindopril, or saline. Six weeks after MI, echocardiography was used to evaluate cardiac structure and function. Myocardial tissue morphology was observed by haematoxylin and eosin (H&E) staining, and heart samples were stained with Masson's trichrome to analyse myocardial fibrosis. Myocardial hypertrophy was observed by fluorescent wheat germ agglutinin (WGA) staining. The expressions of NLRP3, ASC, Cle-caspase-1, IL-1β, TXNIP, Cle-caspase-3, Bcl-2, and Bax in heart tissues were assessed by western blot analysis. mRNA expressions of ANP and BNP in heart tissues were measured by RT-PCR. The expression of reactive oxygen species in myocardial tissue was detected by using a DCFH-DA probe. Results. Echocardiographic analysis showed that compared with the model group, the left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS) in the LQF and perindopril group were increased (P < 0.05), left ventricular internal diameter end diastole (LVIDd) and left ventricular internal diameter end-systole (LVIDs) were reduced (P < 0.05), and H&E and Masson's trichrome staining of cardiac tissues showed that LQF and perindopril could partially reverse ventricular remodeling and alleviate myocardial fibrosis (P < 0.05). WGA fluorescence results showed that compared with the model group, myocardial hypertrophy was significantly reduced in the LQF and perindopril group. We also found that LQF and perindopril reduce the oxidative stress response in the heart of MI mice. The protein expression of NLRP3, ASC, Cle-caspase-1, IL-1β, TXNIP, Cle-caspase-3, and Bax was downregulated in the LHF and perindopril treatment group, and Bcl-2 expression was upregulated. Conclusion. LQF and perindopril significantly attenuated cardiac injury and apoptosis in the MI model. In addition, we found that LQF effectively inhibited the activation of the NLRP3/ASC/caspase-1/IL-1β cascade, decreased inflammatory infiltration, delayed ventricular remodeling, and downregulated caspase-3/Bax signaling, which can effectively reduce the apoptosis of cardiomyocytes. Perindopril showed the same mechanism. [ABSTRACT FROM AUTHOR]

Highlights • A promoter trap system using two reporter genes was developed in P. polymyxa SC2-M1. • Some novel and native promoters of P. polymyxa were identified and characterized. • An XI was expressed in P. polymyxa SC2-M1 to improve the xylose metabolic capacity. Abstract Paenibacillus polymyxa is a rhizobacterium that has attracted substantial attention due to its ability to produce functional metabolites and promote plant growth. Metabolic and genetic improvements in this species will benefit research and other applications of the bacterium. However, a suitable gene expression system has not been established in this species. In this study, a promoter trap system based on a green fluorescent protein and a chloramphenicol-resistance gene was developed to isolate native promoters of P. polymyxa SC2-M1 to regulate gene expression. Through high-throughput screening, the novel promoter P LH-77 was identified, sequenced, and subsequently characterized. Promoter P LH-77 is a strong, continuous expression system containing the typical -10 and -35 motifs regions. Its effective sequence was evaluated and then cascaded to improve the promotion efficiency. To further verify the existence of P LH-77 , a heterogenous xylose isomerase was expressed by P LH-77 in P. polymyxa SC2-M1. In the resulting strain, the amount of xylose consumed was increased by 2.5 g/L during the 78 h fermentation period. Meanwhile, the production levels of lactate and acetate increased. It was confirmed that promoter P LH-77 could effectively mediate gene expression in P. polymyxa SC2-M1 and will further benefit the quantitative monitoring of gene expression in P. polymyxa. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy is a major cause of chronic kidney disease and end-stage kidney disease. However, so little is known about alterations of the glycopatterns in urine with the development of diabetic nephropathy. Presently, we interrogated glycopatterns in urine specimens using a lectin microarray. The results showed that expression levels of Siaα2-6Gal/GalNAc recognized by SNA exhibited significantly increased tendency with the development of diabetic nephropathy; moreover, SNA blotting indicated glycoproteins (90 kDa, 70 kDa, and 40 kDa) in urine may contribute to this alteration. Furthermore, the glycopatterns of (GlcNAc)2–4 recognized by STL exhibited difference between diabetic and nondiabetic nephropathy. The results of urinary protein microarray fabricated by another 48 urine specimens also indicated (GlcNAc)2–4 is a potential indictor to differentiate the patients with diabetic nephropathy from nondiabetic nephropathy. Furtherly, STL blotting showed that the 50 kDa glycoproteins were correlated with this alteration. In conclusion, our data provide pivotal information to monitor the development of diabetic nephropathy and distinguish between diabetic nephropathy and nondiabetic renal disease based on precise alterations of glycopatterns in urinary proteins, but further studies are needed in this regard. [ABSTRACT FROM AUTHOR]