Enzymatic properties and immobilization of a thermostable prenyltransferase from Aspergillus fumigatiaffinis for the production of prenylated naringenin.

Prenyltransferases catalyze the synthesis of prenylated flavonoids, providing these with greater lipid solubility, biological activity, and availability. In this study, a thermostable prenyltransferase (AfPT) from Aspergillus fumigatiaffinis was cloned and expressed in Escherichia coli. By optimizing induction conditions, the expression level of AfPT reached 39.3 mU/mL, which was approximately 200 % of that before optimization. Additionally, we determined the enzymatic properties of AfPT. Subsequently, AfPT was immobilized on carboxymethyl cellulose magnetic nanoparticles (CMN) at a maximum load of 0.6 mg/mg. Optimal activity of CMN-AfPT was achieved at pH 8.0 and 55 °C. Thermostability assays showed that the residual activity of CMN-AfPT was greater than 50 % after incubation at 55 °C for 4 h. Km and Vmax of CMN-AfPT for naringenin were 0.082 mM and 5.57 nmol/min/mg, respectively. The Kcat/Km ratio of CMN-AfPT was higher than that of AfPT. Residual prenyltransferase activity of CMN-AfPT remained higher than 70 % even after 30 days of storage. Further, CMN-AfPT retained 68 % of its original activity after 10 cycles of reuse. Compared with free AfPT, CMN-AfPT showed higher catalytic efficiency, thermostability, metal ion tolerance, substrate affinity, storage stability, and reusability. Our study presents a thermostable prenyltransferase and its immobilized form for the production of prenylated flavonoids in vitro.

Capsaicin ameliorates renal fibrosis by inhibiting TGF-ß1-Smad2/3 signaling.

BACKGROUND AND PURPOSE: Chronic kidney disease (CKD), characterized by renal fibrosis, is a global refractory disease with few effective therapeutic strategies. It has been reported that capsaicin exerts many pharmacological effects including liver and cardiac fibrosis. However, whether capsaicin plays a therapeutic role in renal fibrosis remains unclear. METHODS: We investigated antifibrotic effects of capsaicin in two mouse renal fibrosis models as follows: C57BL/6J mice were subjected to unilateral ureteral obstruction (UUO) and fed with an adenine-rich diet. We uncovered and verified the mechanisms of capsaicin in human proximal tubular epithelial cells (HK2). We mainly used histochemistry, immunohistochemistry and immunofluorescence staining, western blot assay, biochemical examination and other tools to examine the effects of capsaicin on renal fibrosis and the underlying mechanisms. RESULTS: Capsaicin treatment significantly alleviated fibronectin and collagen depositions in the tubulointerstitium of the injured kidneys from UUO and adenine-fed mice. Meanwhile, capsaicin treatment obviously reduced α-SMA expression. Moreover, capsaicin treatment dramatically protected against the phenotypic alteration of tubular epithelial cells by increasing E-cadherin expression and decreasing vimentin expression during renal fibrosis. Mechanistically, capsaicin treatment effectively suppressed α-SMA and vimentin expressions but promoted E-cadherin expression in HK2 cells mainly through the inhibition of TGF-ß1-Smad2/3 signaling. CONCLUSION: Capsaicin significantly ameliorated renal fibrosis possibly by retarding the activation of myofibroblasts and protecting against the phenotypic alteration of tubular epithelial cells mainly through the inhibition of TGF-ß1-Smad2/3 signaling. Thus, our findings may provide a new insight into the clinical application of capsaicin in renal fibrosis.

Template-directed synthesis of pomegranate-shaped zinc oxide@zeolitic imidazolate framework for visible light photocatalytic degradation of tetracycline.

The development of photocatalysts for efficient tetracycline (TC) degradation under visible light is urgently needed yet remains a great challenge. Most semiconductor photocatalysts with low specific surface area are easy to agglomerate in solution and unfavorable for enriching pollutants. Herein, we present the preparation of pomegranate-shaped zinc oxide@zeolitic imidazolate framework (ZnO@ZIF-8) by in situ growth of ZIF-8 on a petal-shaped ZnO template that enhances the adsorption and photocatalytic degradation of TC. ZnO@ZIF-8 exhibits an excellent photostability and a TC photodegradation efficiency of 91% under visible light (λ > 420 nm) in 50 min at room temperature, which can be recycled over five times without any loss of activity. Moreover, the plausible photocatalysis reaction mechanism and the degradation intermediates are elucidated with the aid of three-dimensional excitation-emission matrix spectra and liquid chromatography-mass spectrometry system. This study offers new insights into the design of antibiotic degradation photocatalysts and the development of photocatalysts with broad-spectrum responses for efficient TC elimination.

Increased Circulating PD-1 hi CXCR5 - Peripheral T Helper Cells are Associated with Disease Activity of ANCA-Associated Vasculitis.

Newly identified PD-1 hiCXCR5 -CD4 + T cells, termed as peripheral helper T cells (Tph), have been found elevated and playing pathogenic role in some autoimmune diseases like systemic lupus erythematosus (SLE) and rheumatic arthritis (RA). However, the potential role of Tph cells in Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) remains unclear. Here, we explored the potential clinical significance of circulating Tph cells in the pathogenesis of AAV. Comparing 32 active AAV patients and 18 age- and sex-matched healthy controls (HCs), we found that the frequency of circulating Tph cells was significantly expanded in active AAV patients. Besides, programmed death 1 (PD-1) expression on the surface of Tph cells was significantly up-regulated in active AAV patients. Importantly, the frequency of circulating Tph cells was greatly decreased in AAV patients after receiving treatment. Tph cells frequency was positively correlated with the Birmingham Vasculitis Activity Score (BVAS), C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), neutrophil lymphocyte ratio (NLR) and cellular crescent in active AAV patients, but negatively correlated with fibrosus crescent. Tph cells frequency was also positively correlated with naïve B cells, serum concentration of MPO-ANCAs, serum tumor necrosis factor-α (TNF-α), IL-4, IL-21 and IL-12. However, serum IL-10 exhibited negative correlation with circulating Tph cells in active AAV patients. These results demonstrated that circulating Tph cells are greatly expanded in active AAV patients and are positively associated with serum MPO-ANCAs and disease activity, thus contributing to the pathogenesis of AAV.

Characterisation of the heat shock protein Tid and its involvement in stress response regulation in Apis cerana.

Objective: The impact of various environmental stresses on native Apis cerana cerana fitness has attracted intense attention in China. However, the defence responses of A. cerana cerana to different stressors are poorly understood. Here, we aimed to elucidate the regulatory mechanism mediated by the tumorous imaginal discs (Tid) protein of A. cerana cerana (AccTid) in response to stressors. Methods: We used some bioinformatics softwares to analyse the characterisation of Tid. Then, qRT-PCR, RNA interference and heat resistance detection assays were used to explore the function of Tid in stress response in A. cerana cerana. Results: AccTid is a homologous gene of human Tid1 and Drosophila Tid56, contains a conserved J domain and belongs to the heat shock protein DnaJA subfamily. The level of AccTid induced expression was increased under temperature increases from 40°C to 43°C and 46°C, and AccTid knockdown decreased the heat resistance of A. cerana cerana, indicating that the upregulation of AccTid plays an important role when A. cerana cerana is exposed to heat stress. Interestingly, contrary to the results of heat stress treatment, the transcriptional level of AccTid was inhibited by cold, H2O2 and some agrochemical stresses and showed no significant change under ultraviolet ray and sodium arsenite stress. These results suggested that the requirement of A. cerana cerana for Tid differs markedly under different stress conditions. In addition, knockdown of AccTid increased the mRNA levels of some Hsps and antioxidant genes. The upregulation of these Hsps and antioxidant genes may be a functional complement of AccTid knockdown. Conclusion: AccTid plays a crucial role in A. cerana cerana stress responses and may mediate oxidative damage caused by various stresses. Our findings will offer fundamental knowledge for further investigations of the defence mechanism of A. cerana cerana against environmental stresses.

Clinical significance of C4d deposition in renal tissues from patients with primary Sjögren's syndrome-a preliminary study.

BACKGROUND: To evaluate renal expression of C4d, a complement component in the classical/mannose binding lectin (MBL) pathway, in patients with primary Sjögren's syndrome (pSS)-associated renal impairments. METHODS: We retrospectively reviewed the clinical and pathological data from 39 patients with pSS presenting with renal impairments. C4d was examined in paraffin-embedded biopsy tissues using immunohistochemistry. Glomerular C4d positive was defined when > 75% glomeruli were globally stained. Tubulointerstitial C4d (TI-C4d) were scored semi-quantitatively as 0 (absent), 1 (spotty or weak), 2 (patchy) and 3 (diffuse). A TI-C4d score ≥ 2 was considered TI-C4d positive and included in the TI-C4d+ group and vice versa. Peritubular capillary (PTC) C4d was scored as 0 (absent), 1 (0~10%, minimal), 2 (10%~ 50%, focal), and 3 (> 50%, diffuse). RESULTS: Glomerular C4d deposition was observed in all 8 patients with pSS-related membranous nephropathy (MN) without obvious C1q deposition. Two of 5 patients with mesangial proliferative glomerulonephritis and 1 of 2 patients with IgA nephropathy had mild mesangial C4d deposition. Sixteen patients (6 glomerular dominant and 10 tubulointerstitial dominant) presented TI-C4d score ≥ 2. Patients in the TI-C4d+ group exhibited a higher serum creatinine level at the time of renal biopsy (TI-C4d+ 132.5 [89.7, 165.5] vs. TI-C4d- 83.0 [70.7, 102.0] µmol/L, P = 0.008). PTC C4d was observed in 12 patients, with each of minimal, focal and diffuse staining being noted in 4 patients. CONCLUSIONS: The MBL pathway of complement activation was potentially involved in pSS-related MN. Tubulointerstitial C4d might be a pathological marker of severe renal injury in patients with pSS-related renal impairments.

Congenital hypodysfibrinogenemia associated with a novel deletion of three residues (γAla289_Asp291del) in fibrinogen.

Hypodysfibrinogenemia is the least frequently reported congenital fibrinogen disorder, characterized by both quantity and quality defects of fibrinogen. In this study, we investigated the molecular basis of hypodysfibrinogenemia in a Chinese family. Functional fibrinogen was measured by Clauss method, and the antigenic fibrinogen was measured by immunoturbidimetry assay. All the exons and exon-intron boundaries of fibrinogen genes (FGA, FGB and FGG) were analysed by direct DNA sequencing. To further evaluate its molecular and functional characterizations, fibrinogen was purified from the plasma of propositus, then SDS-PAGE, fibrin polymerization, clot lysis, and electron microscopy scanning were all performed. The propositus showed a slight decrease of immunologic fibrinogen (1.52 g/L) but dramatically reduced functional fibrinogen (0.3 g/L). DNA sequencing revealed a novel heterozygous CCTTTGATG deletion in the exon 8 of FGG, leading to the deletion of Ala289, Phe290, and Asp291 in fibrinogen γ-chain. The polymerization of the fibrinogen from the propositus was markedly impaired, with prolonged lag period and decreased final turbidity. The fibrinogen clottability showed a reduced fraction of participating clot formation. While the clot lysis showed normal. Scanning electron microscopy revealed that the fibers of the propositus were thicker than normal, with larger pores and curlier meshworks. We conclude that γAla289_Asp291del is responsible for the hypodysfibrinogenemia in this case.

A novel mutation (Tyr503Cys) in a severe factor XI deficiency.

: The objective is to study a gene mutation (Tyr503Cys) found in a Chinese consanguineous marriage family with inherited factor XI (FXI) deficiency (cross-reacting material positive, type II). The FXI activity and FXI antigen were tested with clotting assay and ELISA, respectively. The FXI gene was amplified by PCR with direct sequencing. ClustalX-2.1-win and three online bioinformatics softwares were used to study the conservatism and harm of the mutation. The proband had reduced FXI: activity at 13%; three members had decreased to about 35%, all of whom had nomal FXI: antigen. DNA sequencing analysis showed the proband carried a homozygous c.1562A>G point mutation of F11, resulting in Tyr503Cys. Tyr503 was highly conserved among the homologous species. The three bioinformatics softwares indicated that the mutation had affected the function of the protein. The Tyr503Cys mutation was responsible for the decrease of FXI: activity, which is cross-reacting material positive deficiency and the first reported in the world.

Death associated protein 1 (DAP 1) positively regulates virus replication and apoptosis of hemocytes in shrimp Marsupenaeus japonicus.

Death-associated protein 1 (DAP1) is a small proline-rich cytoplasmic protein that functions both in the apoptosis and autophage process of mammalian and in the clinical cancer of human. However, little knowledge is known about the homologue gene of DAP1 and its roles in the physiological process of invertebrates. In this paper, we report a novel function of DAP1 in the antivirus immunity of shrimp. A homologue gene of DAP1 was cloned from Marsupenaeus japonicus and named as Mjdap-1. The full-length of Mjdap-1 was 1761 bp with a 309 bp open reading frame that encoded 102 amino acids. Reverse transcription-PCR results showed that Mjdap-1 was expressed in all tested tissues, including hemocytes, gills, intestines, stomach, heart, hepatopancreas, testes, and ovaries. In shrimp, Mjdap-1 transcripts were up-regulated by white spot syndrome virus (WSSV) infection; Mjdap-1 knockdown decreased the virus copy in vivo and the mortality of M. japonicus to WSSV challenge. Conversely, injecting the purified recombinant MjDAP1 protein promoted the amplification of virus in shrimp. Flow cytometric assay showed, the virus infection-induced apoptosis of hemocytes was enhanced by MjDAP1 protein injection and inhibited in MjDAP1 knockdown shrimp. Furthermore, the expression of apoptosis-inducing factor (AIF) was regulated by Mjdap-1, but the caspase transcripts were not affected. Our results suggested that MjDAP1 facilitated the amplification of virus in shrimp, which may be attributed to the promotion of hemocyte apoptosis in an AIF-dependent manner. These results provided a new insight into the function of this protein that may be used for virus disease control.

Rapidly progressive glomerulonephritis due to anti-glomerular basement membrane disease accompanied by IgA nephropathy: a case report.

Anti-glomerular basement membrane (anti-GBM) disease is characterized by autoantibodies against antigenic site on type IV collagen of the GBM. The coexistence of anti-GBM disease and other immune complex mediated glomerulonephritis is common. Herein, we describe a patient presented with rapidly progressive glomerulonephritis, who was diagnosed as IgA-mediated nephropathy and was found to have abundant serum anti-glomerular basement membrane IgG antibodies. The patient's renal function improved considerably with intensive immunosuppressive therapy.