Antiinflammatory triterpenoids and steroids from Ganoderma lucidum and G. tsugae

The antiinflammatory properties of triterpenoids and steroids from both Ganoderma lucidum and Ganoderma tsugae were studied. Twelve compounds, including ergosta-7,22-dien-3beta-ol (1), ergosta-7,22-dien-3beta-yl palmitate (2), ergosta-7,22-dien-3-one (3), ergosta-7,22-dien-2beta,3alpha,9alpha-triol (4), 5alpha,8alpha-epidioxyergosta-6,22-dien-3beta-ol (5), ganoderal A (6), ganoderal B (7), ganoderic aldehyde A (8), tsugaric acid A (9), 3-oxo-5alpha-lanosta-8,24-dien-21-oic acid (10), 3alpha-acetoxy-5alpha-lanosta-8,24-dien-21-oic acid ester beta-d-glucoside (11), and tsugaric acid B (12), were assessed in vitro by determining their inhibitory effects on the chemical mediators released from mast cells, neutrophils, and macrophages. Compound 10 showed a significant inhibitory effect on the release of beta-glucuronidase from rat neutrophils stimulated with formyl-Met-Leu-Phe (fMLP)/cytochalasin B (CB) whereas compound 9 significantly inhibited superoxide anion formation in fMLP/CB-stimulated rat neutrophils. Compound 10 also exhibited a potent inhibitory effect on NO production in lipopolysaccharide (LPS)/interferon-gamma (IFN-gamma)-stimulated N9 microglial cells. Moreover, compound 9 was also able to protect human keratinocytes against damage induced by ultraviolet B (UV B) light, which indicated 9 could protect keratinocytes from photodamage.     

Anti-obesity properties of a Sasa quelpaertensis extract in high-fat diet-induced obese mice

This study explores the anti-obesity properties of a Sasa quelpaertensis leaf extract (SQE) in high-fat diet (HFD)-induced obese C57BL/6 mice and mature 3T3-L1 adipocytes. SQE administration with HFD for 70 d significantly decreased the body weight gain, adipose tissue weight, and serum total cholesterol and triglyceride levels in comparison with the HFD group. SQE administration also reduced the serum levels of glutamic oxaloacetic transaminase, glutamic pyruvic transaminase and lactate dehydrogenase, and the accumulation of lipid droplets in the liver, suggesting a protective effect against HFD-induced hepatic steatosis. SQE administration restored the HFD-induced decreases with phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) in epididymal adipose tissue. SQE also induced AMPK phosphorylation in mature 3T3-L1 adipocytes. These results suggest that SQE exerted an anti-obesity effect on HFD-induced obese mice by activating AMPK in adipose tissue and reducing lipid droplet accumulation in the liver.     

Silencing or Amplification of Endocannabinoid Signaling in Blastocysts via CB1 Compromises Trophoblast Cell Migration

Endocannabinoid signaling plays key roles in multiple female reproductive events. Previous studies have shown an interesting phenomenon, that mice with either silenced or elevated endocannabinoid signaling via Cnr1 encoding CB(1) show similar defects in several pregnancy events, including preimplantation embryo development. To unravel the downstream signaling of this phenomenon, microarray studies were performed using RNAs collected from WT, Cnr1(-/-), and Faah(-/-) mouse blastocysts on day 4 of pregnancy. The results indicate that about 100 genes show unidirectional changes under either silenced or elevated anandamide signaling via CB(1). Functional enrichment analysis of the microarray data predicted that multiple biological functions and pathways are affected under aberrant endocannabinoid signaling. Among them, genes enriched in cell migration are suppressed in Cnr1(-/-) or Faah(-/-) blastocysts. Cell migration assays validated the prediction of functional enrichment analysis that cell mobility and spreading of either Cnr1(-/-) or Faah(-/-) trophoblast stem cells are compromised. Either silenced or elevated endocannabinoid signaling via CB(1) causes similar changes in downstream targets in preimplantation embryos and trophoblast stem cells. This study provides evidence that a tightly regulated endocannabinoid signaling is critical to normal preimplantation embryo development and migration of trophoblast stem cells.     

Functional and structural studies of pullulanase from Anoxybacillus sp. LM18‐11

Pullulanase is a debranching enzyme that specifically hydrolyzes the α‐1,6 glycosidic linkage of α‐glucans, and has wide industrial applications. Here, we report structural and functional studies of a new thermostable pullulanase from Anoxybacillus sp. LM18‐11 (PulA). Based on the hydrolysis products, PulA was classified as a type I pullulanase. It showed maximum activity at 60°C and pH 6.0. Kinetic study showed that the specific activity and K_m for pullulan of PulA are 750 U mg^?1 and 16.4 μmol L^?1, respectively. PulA has a half‐life of 48 h at 60°C. The remarkable thermostability makes PulA valuable for industrial usage. To further investigate the mechanism of the enzyme, we solved the crystal structures of PulA and its complexes with maltotriose and maltotetraose at 1.75–2.22 Å resolution. The PulA structure comprises four domains (N1, N2, A, and C). A is the catalytic domain, in which three conserved catalytic residues were identified (D413, E442, and D526). Two molecules of oligosaccharides were seen in the catalytic A domain in a parallel binding mode. Interestingly, another two oligosaccharides molecules were found between the N1 domain and the loop between the third β‐strand and the third α‐helix in the A domain. Based on sequence alignment and the ligand binding pattern, the N1 domain is identified as a new type of carbohydrate‐binding motif and classified to the CBM68 family. The structure solved here is the first structure of pullulanase which has carbohydrate bound to the N1 domain. Proteins 2014; 82:1685–1693. © 2013 Wiley Periodicals, Inc.     

The impact of cytomegalovirus infection on clinical severity and outcomes in kidney transplant recipients with Pneumocystis jirovecii pneumonia

Cytomegalovirus (CMV) infection is associated with Pneumocystis jirovecii pneumonia (PJP) in kidney transplant recipients (KTRs), but its impact on clinical severity and outcomes in KTRs with PJP is unknown. We reviewed 1994 medical records of KTRs from January 1997 to March 2019. PJP or CMV infection was diagnosed by polymerase chain reaction or culturing using blood or respiratory specimens. We divided patients into PJP and PJP+CMV groups, and evaluated the clinical severity and outcomes. Fifty two patients had PJP (2.6%) in the whole study cohort. Among patients with PJP, 38 (73.1%) had PJP alone and 14 (26.9%) had combined PJP and CMV co-infection. The PJP+CMV group showed worse laboratory findings (serum albumin and C-reactive protein, P = 0.010 for both) and higher requirement of continuous renal replacement therapy than the PJP group (P = 0.050). The pneumonia severity was worse in the PJP+CMV group than in the PJP group (P < 0.05), and CMV infection was a high risk factor of pneumonia severity (odds ratio 16.0; P = 0.002). The graft function was worse in the PJP+CMV group (P < 0.001), and the incidence of graft failure was higher in the PJP+CMV group than in the PJP group (85.7% vs 36.8%; P < 0.001). Mortality was double in the PJP+CMV group than in the PJP group, but not statistically significant (21.4% vs 10.5%; P = 0.370). Our results show that approximately one in four patients with PJP after kidney transplantation develops CMV with increased clinical severity and risk of graft failure. The possibility of increased clinical severity and worse clinical outcomes by CMV co-infection should be considered in KTRs with PJP.     

PLP2/A4 interacts with CCR1 and stimulates migration of CCR1-expressing HOS cells

Multiple CC chemokines bind to CCR1, which plays important roles in immune and inflammatory responses. To search for proteins involved in the CCR1 signaling pathway, we screened a yeast two-hybrid library using the cytoplasmic tail of CCR1 as the bait. One of the positive clones contained an open reading frame of 456bp, of which the nucleotide sequence was identical to that of proteolipid protein 2 (PLP2), also known as protein A4. Mammalian two-hybrid and coimmunoprecipitation analyses demonstrated the association of PLP2/A4 with CCR1. Indirect immunofluorescence analysis revealed that PLP2/A4 was predominantly located in plasma membrane and colocalized with CCR1 in transfected human HEK293 cells. In addition, focal staining of CCR1 appeared on the periphery of the membrane upon short exposure to Leukotactin-1(Lkn-1)/CCL15, a CCR1 agonist, and was costained with PLP2/A4 on the focal regions. PLP2/A4 mRNAs were detected in various cells such as U-937, HL-60, HEK293, and HOS cells. Overexpression of PLP2/A4 stimulated a twofold increase in the agonist-induced migration of HOS/CCR1 cells, implicating a functional role for PLP2/A4 in the chemotactic processes via CCR1.