Beneficial Effects of Echinacoside on Cognitive Impairment and Diabetes in Type 2 Diabetic db/db Mice.

INTRODUCTION: Cognitive dysfunction is an important comorbidity of diabetes. Insulin resistance may play a critical role in diabetes-related cognitive impairment. Echinacoside (ECH), a natural phenylethanoid glycoside, is the active component of anti-diabetes prescriptions in traditional Chinese medicine. Its effect on modulating insulin resistance has been confirmed but modulating neurodegenerative disease remains unclear. METHODS: Db/db mice, a spontaneous type 2 diabetes mode, were intragastrically administered ECH by 300 mg/kg or an equivalent volume of saline. Weight, blood glucose, and insulin resistance index were measured. Morris water maze test was performed to observe the compound effects on cognition. Hippocampal lesions were observed by histochemical analysis. RESULTS: In db/db mice, ECH alleviated diabetes symptoms, memory loss, and hippocampal neuronal damage. Next, the expression of CD44 and phosphorylated tau was upregulated in diabetic mice. In addition, the insulin receptor substrate-1/phosphatidylinositol 3-kinase /protein kinase B signaling pathway was dysregulated in diabetic mice. All these dysregulations could be reversed by ECH. DISCUSSION: This study provides theoretical support and experimental evidence for the future application of ECH in diabetic cognition dysfunction treatment, promoting the development of traditional medicines.

Molecular and functional characterization of Schistosoma japonicum annexin A13.

Schistosomiasis is a neglected tropical disease that affects humans and animals in tropical and subtropical regions worldwide. Schistosome eggs are responsible for the pathogenesis and transmission of schistosomiasis, thus reducing egg production is vital for prevention and control of schistosomiasis. However, the mechanisms underlying schistosome reproduction remain unclear. Annexin proteins (ANXs) are involved in the physiological and pathological functions of schistosomes, but the specific regulatory mechanisms and roles of ANX A13 in the development of Schistosoma japonicum and host-parasite interactions remain poorly understood. Therefore, in this study, the expression profiles of SjANX A13 at different life cycle stages of S. japonicum were assessed using quantitative PCR. In addition, the expression profiles of the homolog in S. mansoni were analyzed in reference to public datasets. The results of RNA interference showed that knockdown of SjANX A13 significantly affected the development and egg production of female worms in vivo. The results of an immune protection assay showed that recombinant SjANX A13 increased production of immunoglobulin G-specific antibodies. Finally, co-culture of S. japonicum exosomes with LX-2 cells using a transwell system demonstrated that SjANX A13 is involved in host-parasite interactions via exosomes. Collectively, these results will help to clarify the roles of SjANX A13 in the development of S. japonicum and host-parasite interactions as a potential vaccine candidate.

Functional characterization of differentially expressed proteins coming from unisexual and bisexual infected Schistosoma japonicum female worms.

Schistosomiasis is an important zoonotic disease affecting up to 40 kinds of animals and is responsible for ∼250 million human cases per year. Due to the extensive use of praziquantel for the treatment of parasitic diseases, drug resistance has been reported. Consequently, novel drugs and effective vaccines are urgently needed for sustained control of schistosomiasis. Targeting reproductive development of Schistosoma japonicum could contribute to the control of schistosomiasis. In this study, five highly expressed proteins (S. japonicum large subunit ribosomal protein L7e, S. japonicum glutathione S-transferase class-mu 26 kDa isozyme, S. japonicum UDP-galactose-4-epimerase and two hypothetical proteins SjCAX70849 and SjCAX72486) in 18, 21, 23, and 25-day mature female worms compared to single-sex infected female worms were selected based on our previous proteomic analysis. Quantitative real-time polymerase chain reaction analysis and long-term interference with small interfering RNA were performed to identify the biological functions of these five proteins. The transcriptional profiles suggested that all five proteins participated in the maturation of S. japonicum. RNA interference against these proteins resulted in morphological changes to S. japonicum. The results of an immunoprotection assay revealed that immunization of mice with recombinant SjUL-30 and SjCAX72486 upregulated production of immunoglobulin G-specific antibodies. Collectively, the results demonstrated that these five differentially expressed proteins were vital to reproduction of S. japonicum and, thus, are potential candidate antigens for immune protection against schistosomiasis.

Comparative proteomic profiles of Schistosoma japonicum male worms derived from single-sex and bisexual infections.

Schistosomiasis, which is caused by parasitic schistosomes, remains the second most prevalent parasitic disease of mammals worldwide. To successfully maintain fecundity, schistosomes have evolved a lifecycle that involves the cooperation of morphologically distinct male and female forms. Eggs produced by worm pairs are vital to the lifecycle of the parasite and are responsible for pathogenesis. Understanding the reproductive mechanism of schistosomes will help to control infection. In this study, the proteomic profiles of single-sex infected male (SM) worms and bisexual infected mated male (MM) worms of Schistosoma japonicum at 18, 21, 23, and 25 days p.i. were identified through data-independent acquisition. In total, 674 differentially expressed proteins (DEPs) were identified for the SM and MM worms at all four timepoints. Bioinformatic analysis demonstrated that most of the DEPs were involved in biosynthetic processes including locomotion, cell growth and death, cell motility, and metabolic processes such as protein metabolism and glucose metabolism. Schistosoma japonicum glycosyltransferase (SjGT) and S. japonicum nicastrin protein (SjNCSTN) were selected for quantitative real­time PCR analysis and long-term interference with small interfering RNA (siRNA) to further explore the functions of the DEPs. Sjgt mRNA expression was mainly enriched in male worms, while Sjncstn was enriched in both sexes. siRNA against SjGT and SjNCSTN resulted in minor morphological changes in the testes of male worms and significant decreased vitality and fertility. The present study provides comprehensive proteomic profiles of S. japonicum SM and MM worms at 18, 21, 23, and 25 days p.i. and offers insights into the mechanisms underlying the growth and maturation of schistosomes.

Schistosoma japonicum translationally controlled tumour protein, which is associated with the development of female worms, as a target for control of schistosomiasis.

Schistosomiasis is a globally important helminthic disease of both humans and animals, and is the second most common parasitic disease after malaria. Although praziquantel is extensively used for treatment of parasitic diseases, drug resistance has been reported. Therefore, new drugs and effective vaccines are needed for continuous control of schistosomiasis. Eggs produced by schistosomes are responsible for the occurrence and spread of schistosomiasis. Revealing the reproductive mechanism of schistosomes will help to control this disease. In this study, the proteomic profiles of single-sex infected female worms and bisexual infected mature female worms of Schistosoma japonicum at 18, 21, 23 and 25 days p.i. were identified with isobaric tags for relative quantitation-coupled liquid chromatography-tandem mass spectrometry. Differentially expressed proteins were subsequently used for bioinformatic analysis. Six highly expressed differentially expressed proteins in mature female worms were selected and long-term interference with small interfering RNA (siRNA) was conducted to determine biological functions. SiRNA against S. japonicum translationally controlled tumour protein (SjTCTP) resulted in the most significant effect on the growth and development of MF worms. Sjtctp mRNA expression gradually increased over time with a high level of expression maintained at 25-42 days p.i., while levels were significantly higher in mature female worms than male and SF worms. The subsequent animal immune protection experiments showed that recombinant SjTCTP (rSjTCTP) reduced the number of adults by 44.7% (P < 0.01), average egg burden per gram of liver by 57.94% (P < 0.01), egg hatching rate by 47.57% (P < 0.01), and oviposition of individual females by 43.16%. rSjTCTP induced higher levels of serum IgG, IL-2, and IL-10 in mice. Collectively, these results show that SjTCTP is vital to reproduction of female worms and, thus, is a candidate antigen for immune protection.

Sensitivity to Morphine Reward Associates With Gut Dysbiosis in Rats With Morphine-Induced Conditioned Place Preference.

Gut microbiota has been found to establish a bidirectional relationship with the central nervous system. Variations of the gut microbiota has been implicated in various mental disorders, including opioid use disorders. Morphine exposure has been repeatedly found to disrupt the gut microbiota, but association between the gut microbiota and the sensitivity to morphine reward remains unknown. In this study the conditioned place preference (CPP) paradigm was used for morphine-treated rats and saline-treated rats. After the CPP procedure, the morphine-treated rats were divided equally into the low and high CPP (L- and H-CPP) groups according to the CPP scores. We adopted 16S rRNA sequencing for the fecal bacterial communities at baseline and post-conditioning. By comparing the morphine-treated group with saline-treated group, we found alterations of microbial composition in the morphine-treated group, but no significant differences in alpha diversity. The L-CPP group and H-CPP group differed in microbial composition both before and after morphine treatment. The relative abundance of certain taxa was correlated to the CPP scores, such as Alloprevotella and Romboutsia. This study provides direct evidence that morphine exposure alters the composition of the gut microbiota in rats and that microbial alterations are correlated to the sensitivity to morphine reward. These findings may help develop novel therapeutic and preventive strategies for opioid use disorder.

Comparative analysis of iTRAQ-based proteome profiles of Schistosoma japonicum female worms coming from single-sex infections and bisexual infections.

In schistosomiasis, eggs produced by bisexual infected mature female schistosome worms (FMS) are the main cause of pathological damage to the host and the dissemination of the disease. Single-sex infected female worms (FSS) cannot completely develop to sexual maturity or produce normal eggs. In this study, isobaric tags for relative and absolute quantitation (iTRAQ)-coupled LC-MS/MS was used to explore the proteome of FSS and FMS of Schistosoma japonicum. A total of 1477 differentially expressed proteins (fold change >1.2, P < .05) between FSS and FMS were identified. Bioinformatics analysis indicated that FMS expressed more proteins related to biosynthetic processes, such as eggshell synthesis, ribosomal synthesis, protein folding, cellular detoxification, and metabolic processes such as protein metabolism and glucose metabolism, whereas more proteins related to locomotion and oxidative phosphorylation were expressed in FSS. Our identification and analysis of differentially expressed proteins between FMS and FSS provides new insights to elucidate the molecular biological mechanisms of female worm sexual maturation and reproduction. SIGNIFICANCE: Female Schistosome worms must maintain constant pairing contact with male worms for differentiation of their reproductive organs. Mature female worms can produce infectious eggs, cause serious pathological damage to the host and the dissemination of the disease. Unpaired female worms remain small and sexually immature; they do not spawn normally. In this study, iTRAQ-coupled LC-MS/MS was used to explore the whole proteome of single-sex infected female worms (FSS) and bisexual infected mature female worms (FMS) of Schistosoma japonicum. 1477 differentially expressed proteins (DEPs) between FSS and FMS were identified and analyzed. Further research on DEPs' functions in schistosome sexual maturation and reproductive development might provide theoretical bases to explore female maturation and spawning.

Functional analysis of the Frzb2 gene in Schistosoma japonicum.

Schistosomiasis is a globally important helminthic disease of humans and animals, and it is the second most common parasitic disease after malaria. Eggs produced by mature females are responsible for the disease's occurrence and spread. Frzb2, a secreted frizzled-related protein, can inhibit Wnt signalling by competitive binding to the specific frizzled protein receptor. In this study, the complete gene sequence of SjFrzb2 was obtained by using 3'-rapid amplification of cDNA ends technology. SjFrzb2 transcript levels at different stages of S. japonicum maturation were evaluated by quantitative real-time RT-PCR analysis. SjFrzb2 was expressed at all developmental stages examined and exhibited the highest transcription level in 7-day-old worms, then gradually decreased during the growth and developmental stages to reach the lowest level at 18 days post-infection. SjFrzb2 gene expression was higher in female worms than in male worms and was significantly higher in female worms from a single-sex infection than in female worms from a bisexual infection. The functions of SjFrzb2 were explored via a small interfering RNA-based gene silencing approach and the soaking method. The results showed that SjFrzb2 gene knockdown impaired the growth and development of S. japonicum in mice, affecting not only the survival and morphological structure of the worms but also their reproductive ability and the viability of the produced eggs. Collectively, these observations imply that Frzb2 may be a novel target for the development of immuno- and/or small molecule-based therapeutics to control schistosomiasis fecundity and transmission.

Function of the lesswright (lwr) gene in the growth, development, and reproduction of Schistosoma japonicum.

The lesswright (lwr) gene and its products are essential molecules in mitosis, DNA repair, and embryo formation in many eukaryotes. In this study, immunohistochemical analysis revealed that the Lwr protein was located in the internal tissues and the surface layer of the adult Schistosoma japonicum (Sj) worms. The mRNA expression levels of SjLwr at different points were evaluated by quantitative real-time RT-PCR. The expression of SjLwr peaked at 14 days and then decreased thereafter. SjLwr expression was relatively more stable in male worms than in female worms. The functions of SjLwr were explored by siRNA-based gene silencing with a simple soaking method. The results showed that knockdown of the SjLwr gene impaired the growth and development of S. japonicum in mice, as well as survival, morphology, reproductive capacity, and egg vitality. These observations imply that SjLwr presents a novel target for the development of immuno- and/or small molecule-based therapeutics for the control and treatment of schistosome infections.