A Novel PEGylated Liposome-Encapsulated SANT75 Suppresses Tumor Growth through Inhibiting Hedgehog Signaling Pathway

The Hedgehog (Hh) pathway inhibitors have shown great promise in cancer therapeutics. SANT75, a novel compound we previously designed to specially inhibit the Smoothened (SMO) protein in the Hh pathway, has greater inhibitory potency than many of commonly used Hh inhibitors. However, preclinical studies of SANT75 revealed water insolubility and acute toxicity. To overcome these limitations, we developed a liposomal formulation of SANT75 and investigated its antitumor efficacy in vitro and in vivo. We encapsulated SANT75 into PEGylated liposome and the mean particle size distribution and zeta-potential (ZP) of liposomes were optimized. Using the Shh-light2 cell and Gli-GFP or Flk-GFP transgenic reporter zebrafish, we confirmed that liposome-encapsulated SANT75 inhibited Hh activity with similar potency as the original SANT75. SANT75 encapsulated into liposome exerted strong tumor growth-inhibiting effects in vitro and in vivo. In addition, the liposomal SANT75 therapy efficiently improved the survival time of tumor-bearing mice without obvious systemic toxicity. The pathological morphology and immunohistochemistry staining revealed that liposomal SANT75 induced tumor cell apoptosis, inhibited tumor angiogenesis as assessed by CD31 and down-regulated the expression of Hh target protein Gli-1 in tumor tissues. Our findings suggest that liposomal formulated SANT75 has improved solubility and bioavailability and should be further developed as a drug candidate for treating tumors with abnormally high Hh activity.     

Determination of Peripheral Neuropathy Prevalence and Associated Factors in Chinese Subjects with Diabetes and Pre-Diabetes – ShangHai Diabetic neuRopathy Epidemiology and Molecular Genetics Study (SH-DREAMS)

Objective

This study determined the prevalence and factors associated with peripheral neuropathy (PN) in subjects with diabetes mellitus, impaired glucose regulation (IGR), and normal glucose tolerance (NGT) in a community-based Chinese population.

Research Design and Methods

A total of 2035 subjects in Shanghai were classified as having NGT, IGR, or diabetes. All subjects underwent complete foot examination. PN was assessed according to the neuropathy symptom and neuropathy disability scores. Binary logistic regression was performed to analyze the contributions of factors to PN.

Results

The prevalence of PN was 8.4%, 2.8%, and 1.5% in diabetes mellitus, IGR, and NGT subjects, respectively (P<0.05 for diabetes vs. NGT, and IGR). The subjects with known diabetes had the highest frequency of PN (13.1%). Among the subjects without diabetes, those with PN were older, had a higher waist circumference and 2-h postprandial plasma glucose levels, and were more likely to be hypertensive. Among the IGR subjects, other than age, the 2-h postprandial plasma glucose level was an independent factor significantly associated with PN. Meanwhile, among the subjects with diabetes, PN was associated with fasting plasma glucose, duration of diabetes, and decreased estimated glomerular filtration rate.

Conclusions

The prevalence of PN is slightly higher in individuals with IGR than that in individuals with NGT, but small fibre damage in IGR as the earliest nerve fibre deficit may be underestimated in our study. As an independent risk factor, postprandial plasma glucose level may be an important target for strategies to prevent or improve PN in IGR subjects.     

Predictive power of splenic thickness for post-hepatectomy liver failure in HBV-associated hepatocellular carcinoma patients

Background

The purpose of this case series is to investigate the relationship between splenic thickness (ST) and postoperative outcomes after hepatic resection in hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) patients.

Methods

The clinical data of 320 patients with HBV-associated HCC who had undergone liver resection were retrospectively analyzed. The value of ST in predicting postoperative outcomes was evaluated.

Results

A total of 320 patients were enrolled in the study. An increase in ST was significantly associated with an increase in portal vein diameter (PVD), indocyanine green retention rate 15 min (ICG R15), and total bilirubin (TBIL); however, it was negatively correlated with platelet count (PLT). Post-hepatectomy liver failure (PHLF) occurred in 35 (10.9%) patients. Multivariate logistic regression analysis showed that ST was an independent predictor of morbidity and mortality after hepatectomy. Meanwhile, ST was associated with an almost sixfold increased risk for developing perioperative complications (OR 5.678; 95% CI 2.873 to 11.224; P?<?0.001) and almost 13-fold increased risk for mortality after hepatectomy (OR 13.007; 95% CI 1.238 to 136.627; P?=?0.033).The area under the receiver operating characteristic (ROC) curve (AUC) of ST for predicting the incidence of PHLF was 0.754 (95% confidence interval (CI) 0.667 to 0.841; P?<?0.001), with a sensitivity of 57.1% and a specificity of 82.5%, which were significantly greater than those of the ICG R15 level (AUC 0.670; 95% CI 0.560 to 0.779; P?<?0.001). The critical value of ST was 43.5 mm.

Conclusions

ST, which is an easy, inexpensive, and routinely available perioperative marker, showed a favorable predictive value for postoperative outcomes in HBV-associated HCC patients.

     

Effect of culture condition on the growth,biochemical composition and EPA production of alkaliphilic Nitzschia plea isolated in the Southeast of China

Bioprocess and Biosystems Engineering - To overcome the contamination in open pond, microalgal strain selection should focus on species with tolerability to extreme environments. In this study, a...     

Propofol Prevents Oxidative Stress by Decreasing the Ischemic Accumulation of Succinate in Focal Cerebral Ischemia–Reperfusion Injury

Oxidative stress caused by mitochondrial dysfunction during reperfusion is a key pathogenic mechanism in cerebral ischemia–reperfusion (IR) injury. Propofol (2,6-diisopropylphenol) has been proven to attenuate mitochondrial dysfunction and reperfusion injury. The current study reveals that propofol decreases oxidative stress injury by preventing succinate accumulation in focal cerebral IR injury. We evaluated whether propofol could attenuate ischemic accumulation of succinate in transient middle cerebral artery occlusion in vivo. By isolating mitochondria from cortical tissue, we also examined the in vitro effects of propofol on succinate dehydrogenase (SDH) activity and various mitochondrial bioenergetic parameters related to oxidative stress injury, such as the production of reactive oxidative species, membrane potential, Ca²⁺-induced mitochondrial swelling, and morphology via electron microscopy. Propofol significantly decreased the ischemic accumulation of succinate by inhibiting SDH activity and inhibited the oxidation of succinate in mitochondria. Propofol can decrease membrane potential in normal mitochondria but not in ischemic mitochondria. Propofol prevents Ca²⁺-induced mitochondrial swelling and ultrastructural changes to mitochondria. The protective effect of propofol appears to act, at least in part, by limiting oxidative stress injury by preventing the ischemic accumulation of succinate.     

Cloning and functional analysis of PKZ (PKR-like) from grass carp (Ctenopharyngodon idellus)

The new teleost fish PKZ (PKR-like) full-length cDNA (GU299765) had been cloned and identified from grass carp (Ctenopharyngodon idellus). The cDNA of grass carp PKZ (CiPKZ) has 2185 bp in length with a largest open reading frame (ORF) encoding 513aa. CiPKZ possesses a conserved C-terminal catalytic domain of eIF2α kinase family. Within its N-terminal there are two binding domain (Zα) named Zα1 (1-67aa) and Zα2 (81-152aa). BLAST homologous search reveals that CiPKZ has a high-level homology with other fish PKZs and PKRs. Like other fish PKZs and PKRs, CiPKZ is a ubiquitous tissue expression gene that had a very low level of constitutive expression but up-regulated in response to Poly I:C or hot stress (34 °C). For the purpose of searching for the potential function of CiPKZ, we obtained CiPKZ polypeptide via Escherichia coli Rosetta prokaryotic expression and purified with Ni-NTA His-Bind Resin affinity chromatography. CiPKZ polypeptide was used for the test of phosphorylating eIF2αin vitro. The results demonstrated that CiPKZ could be activated by Z-DNA but not by Poly I:C, and with subsequent could phosphorylate eIF2α. Meanwhile, four pcDNA3.1/PKZ recombinant plasmids, including pcDNA3.1/PKZ-wet, pcDNA3.1/PKZ-wet-K198R, pcDNA3.1/PKZ-wet-C, pcDNA3.1/PKZ-wet-C-K198R had been constructed, respectively. Mouse Myeloma cells (Sp2/0) and Human Umbilical Vein Endothelial Cells (HUVEC) were transiently cotransfected with pcDNA3.1/PKZ recombinant plasmid and PGL-3-promoter plasmid. The results revealed that CiPKZ could greatly decrease luciferase level in these cells. Zα and the K198 amino acid residue may play a key role in its function.     

Patterns and processes in the genetic differentiation of the Brachionus calyciflorus complex, a passively dispersing freshwater zooplankton

Elucidating the evolutionary patterns and processes of extant species is an important objective of any research program that seeks to understand population divergence and, ultimately, speciation. The island-like nature and temporal fluctuation of limnetic habitats create opportunities for genetic differentiation in rotifers through space and time. To gain further understanding of spatio-temporal patterns of genetic differentiation in rotifers other than the well-studied Brachionus plicatilis complex in brackish water, a total of 318 nrDNA ITS sequences from the B. calyciflorus complex in freshwater were analysed using phylogenetic and phylogeographic methods. DNA taxonomy conducted by both the sequence divergence and the GMYC model suggested the occurrence of six potential cryptic species, supported also by reproductive isolation among the tested lineages. The significant genetic differentiation and non-significant correlation between geographic and genetic distances existed in the most abundant cryptic species, BcI-W and Bc-SW. The large proportion of genetic variability for cryptic species Bc-SW was due to differences between sampling localities within seasons, rather than between different seasons. Nested Clade Analysis suggested allopatric or past fragmentation, contiguous range expansion and long-distance colonization possibly coupled with subsequent fragmentation as the probable main forces shaping the present-day phylogeographic structure of the B. calyciflorus species complex.     

Isoflurane-induced spatial memory impairment in mice is prevented by the acetylcholinesterase inhibitor donepezil

Although many studies have shown that isoflurane exposure impairs spatial memory in aged animals, there are no clinical treatments available to prevent this memory deficit. The anticholinergic properties of volatile anesthetics are a biologically plausible cause of cognitive dysfunction in elderly subjects. We hypothesized that pretreatment with the acetylcholinesterase inhibitor donepezil, which has been approved by the Food and Drug Administration (FDA) for the treatment of Alzheimer's disease, prevents isoflurane-induced spatial memory impairment in aged mice. In present study, eighteen-month-old mice were administered donepezil (5 mg/kg) or an equal volume of saline by oral gavage with a feeding needle for four weeks. Then the mice were exposed to isoflurane (1.2%) for six hours. Two weeks later, mice were subjected to the Morris water maze to examine the impairment of spatial memory after exposure to isoflurane. After the behavioral test, the mice were sacrificed, and the protein expression level of acetylcholinesterase (AChE), choline acetylase (ChAT) and α7 nicotinic receptor (α7-nAChR) were measured in the brain. Each group consisted of 12 mice. We found that isoflurane exposure for six hours impaired the spatial memory of the mice. Compared with the control group, isoflurane exposure dramatically decreased the protein level of ChAT, but not AChE or α7-nAChR. Donepezil prevented isoflurane-induced spatial memory impairments and increased ChAT levels, which were downregulated by isoflurane. In conclusions, pretreatment with the AChE inhibitor donepezil prevented isoflurane-induced spatial memory impairment in aged mice. The mechanism was associated with the upregulation of ChAT, which was decreased by isoflurane.     

锰作用下PC12细胞的氧化应激机制研究

目的：以鼠嗜铬神经瘤细胞（PC12）为模型,筛选锰对神经细胞增殖抑制作用的时间及剂量,观察锰作用下PC12细胞的氧化应激反应与细胞形态学、生化指标改变和丝裂原活化蛋白激酶pp38（p38MAPKs）的活化表达。方法：用200,400,600,800μmol/LMnCl2的培养液,分别作用对数生长期PC12细胞1,2,3,4d后,用MTT筛选锰的细胞毒性剂量;测定200-600μmol/L MnCl2作用4d后,PC12细胞还原型谷胱甘肽和丙二醛含量;透射电镜观察细胞形态学变化;琼脂糖凝胶电泳检测MnCl2对PC12细胞基因组DNA的影响。western-blot法检测p-p38。结果：MTT实验显示200~800μmol/LMnCl2作用1,2,3,4d对PC12有显著的抑制作用,呈剂量和时间依赖趋势,600μmol/LMnCl2作用4d对PC12的抑制率可达50%以上。200-600μmol/LMnCl2作用于细胞4d后,随着浓度的升高,还原型GSH逐渐降低,MDA的含量逐渐升高;600μmol/LMnCl2作用4d电镜可见细胞凋亡,同样条件下细胞DNA碎片化。Western-blot实验显示600μmol/LMnCl2作用1,2,3,4dp-p38逐渐升高,3d时较对照组增加6.6倍（n=3,p〈0.05）,200,400,600μmol/L MnCl2作用4d时,磷酸化蛋白38（p-p38）也逐渐升高,400μmol/L MnCl2作用4d时较对照组升高了4.7倍（n=3,p〈0.05）。结论：PC12细胞在锰作用下发生氧化应激反应,上调p-p38,诱导细胞凋亡,细胞增殖抑制。