Immunodetection of urinary C-terminal telopeptide fragment of type II collagen: An osteoarthritis biomarker analysis.

The urinary C-terminal telopeptide fragment of type II collagen (uCTX-II) has been reported as the efficient blood-based biomarker for osteoarthritis, which affects knees, hands, spine, and hips. This study reports a sensing strategy with antibody-conjugated gold nanoparticles (GNP) on an interdigitated electrode (IDE) to determine uCTX-II. The GNP-antibody complex was chemically immobilized on the IDE surface through the amine linker. uCTX-II was determined by monitoring the alteration in current upon interacting the GNP-complexed antibody. This strategy was improved the detection by attracting higher uCTX-II molecules, and the detection limit falls in the range of 10-100 pM with an acceptable regression value [y = 0.6254x - 0.4073, R² = 0.9787]. The sensitivity of the detection was recognized at 10 pM. Additionally, upon increasing the uCTX-II concentration, the current changes were increased in a linear fashion. Control detection with nonimmune antibody and control protein do not increase the current level, confirming the specific detection of uCTX-II. This method of detection helps in diagnosing osteoarthritis and its follow-up treatment.

Plasmodium vivax HAP2/GCS1 gene exhibits limited genetic diversity among parasite isolates from the Greater Mekong Subregion.

BACKGROUND: Antigens expressed in sexual stages of the malaria parasites are targets of transmission-blocking vaccines (TBVs). HAP2/GCS1, a TBV candidate, is critical for fertilization in Plasmodium. Here, the genetic diversity of PvHAP2 was studied in Plasmodium vivax parasite populations from the Greater Mekong Subregion (GMS). METHODS: Plasmodium vivax clinical isolates were collected in clinics from the China-Myanmar border region (135 samples), western Thailand (41 samples) and western Myanmar (51 samples). Near full-length Pvhap2 (nucleotides 13-2574) was amplified and sequenced from these isolates. Molecular evolution studies were conducted to evaluate the genetic diversity, selection and population differentiation. RESULTS: Sequencing of the pvhap2 gene for a total of 227 samples from the three P. vivax populations revealed limited genetic diversity of this gene in the GMS (π = 0.00036 ± 0.00003), with the highest π value observed in Myanmar (0.00053 ± 0.00009). Y133S was the dominant mutation in the China-Myanmar border (99.26%), Myanmar (100%) and Thailand (95.12%). Results of all neutrality tests were negative for all the three populations, suggesting the possible action of purifying selection. Codon-based tests identified specific codons which are under purifying or positive selections. Wright's fixation index showed low to moderate genetic differentiation of P. vivax populations in the GMS, with FST ranging from 0.04077 to 0.24833, whereas high levels of genetic differentiation were detected between the China-Myanmar border and Iran populations (FST = 0.60266), and between Thailand and Iran populations (FST = 0.44161). A total of 20 haplotypes were identified, with H2 being the abundant haplotype in China-Myanmar border, Myanmar and Thailand populations. Epitope mapping prediction of Pvhap2 antigen showed that high-score B-cell epitopes are located in the S307-G324, L429-P453 and V623-D637 regions. The E317K and D637N mutations located within S307-G324 and V623-D637 epitopes slightly reduced the predicted score for potential epitopes. CONCLUSIONS: The present study showed a very low level of genetic diversity of pvhap2 gene among P. vivax populations in the Greater Mekong Subregion. The relative conservation of pvhap2 supports further evaluation of a Pvhap2-based TBV.

Bizarreness of Lucid and Non-lucid Dream: Effects of Metacognition.

Dreams are usually characterized by primary consciousness, bizarreness and cognitive deficits, lacking metacognition. However, lucid dreaming (LD) is a type of consciousness state during which the dreamer is aware of the fact that he or she is dreaming, without leaving the sleeping state. Brain research has found that LD shares some common neural mechanisms with metacognition such as self-reflection. With a different metacognition level, the bizarreness of LD would also change. However, the difference in bizarreness between LD and non-LD was seldom explored, and individual differences were often neglected. In the present study, considering LD prevalence in Asia was rarely studied and related results in China and Japan were very different from each other, we first investigated the LD frequency of China in a standardized way. On that basis, we collected dreams of subjects who had relatively higher LD frequency and compared bizarreness density (BD) of LD and non-LD. Moreover, to explore the relationships of metacognition traits and BD, we also measured self-reflection and insight trait by Self-Reflection and Insight Scale. We found that 81.3% of subjects have experienced LD once or more, which is similar to findings in some western countries. Besides, BD was significantly lower in LD than in non-LD. Self-reflection and insight were inversely associated with dream bizarreness. These findings indicate that self-consciousness traits extend from waking to LD and non-LD state. As a particular consciousness state, LD may shed light on the research of consciousness and dream continuity. Future research on dream bizarreness is suggested to take dream types and metacognition differences into consideration.

Genetic diversity, natural selection and haplotype grouping of Plasmodium vivax Duffy-binding protein genes from eastern and western Myanmar borders.

BACKGROUND: Merozoite proteins of the malaria parasites involved in the invasion of red blood cells are selected by host immunity and their diversity is greatly influenced by changes in malaria epidemiology. In the Greater Mekong Subregion (GMS), malaria transmission is concentrated along the international borders and there have been major changes in malaria epidemiology with Plasmodium vivax becoming the dominant species in many regions. Here, we aimed to evaluate the genetic diversity of P. vivax Duffy-binding protein gene domain II (pvdbp-II) in isolates from the eastern and western borders of Myanmar, and compared it with that from global P. vivax populations. METHODS: pvdbp-II sequences were obtained from 85 and 82 clinical P. vivax isolates from the eastern and western Myanmar borders, respectively. In addition, 504 pvdbp-II sequences from nine P. vivax populations of the world were retrieved from GenBank and used for comparative analysis of genetic diversity, recombination and population structure of the parasite population. RESULTS: The nucleotide diversity of the pvdbp-II sequences from the Myanmar border parasite isolates was not uniform, with the highest diversity located between nucleotides 1078 and 1332. Western Myanmar isolates had a unique R391C mutation. Evidence of positive natural selection was detected in pvdbp-II gene in P. vivax isolates from the eastern Myanmar area. P. vivax parasite populations in the GMS, including those from the eastern, western, and central Myanmar as well as Thailand showed low-level genetic differentiation (FST, 0.000-0.099). Population genetic structure analysis of the pvdbp-II sequences showed a division of the GMS populations into four genetic clusters. A total of 60 PvDBP-II haplotypes were identified in 210 sequences from the GMS populations. Among the epitopes in PvDBP-II, high genetic diversity was found in epitopes 45 (379-SIFGT(D/G)(E/K)(K/N)AQQ(R/H)(R/C)KQ-393, π = 0.029) and Ia (416-G(N/K)F(I/M)WICK(L/I)-424], Ib [482-KSYD(Q/E)WITR-490, π = 0.028) in P. vivax populations from the eastern and western borders of Myanmar. CONCLUSIONS: The pvdbp-II gene is genetically diverse in the eastern and western Myanmar border P. vivax populations. Positive natural selection and recombination occurred in pvdbp-II gene. Low-level genetic differentiation was identified, suggesting extensive gene flow of the P. vivax populations in the GMS. These results can help understand the evolution of the P. vivax populations in the course of regional malaria elimination and guide the design of PvDBP-II-based vaccine.

An MFS-Domain Protein Pb115 Plays a Critical Role in Gamete Fertilization of the Malaria Parasite Plasmodium berghei.

Sexual reproduction is an essential process in the Plasmodium life cycle and a vulnerable step for blocking transmission from the human host to mosquitoes. In this study, we characterized the functions of a conserved cell membrane protein P115 in the rodent malaria parasite Plasmodium berghei ANKA. Pb115 was expressed in both asexual stages (schizonts) and sexual stages (gametocytes, gametes, and ookinetes), and was localized on the plasma membrane of gametes and ookinetes. In P. berghei, genetic deletion of Pb115 (Δpb115) did not affect asexual multiplication, nor did it affect gametocyte development or exflagellation of the male gametocytes. However, mosquitoes fed on Δpb115-infected mice showed 74% reduction in the prevalence of infection and 96.5% reduction in oocyst density compared to those fed on wild-type P. berghei-infected mice. The Δpb115 parasites showed significant defects in the interactions between the male and female gametes, and as a result, very few zygotes were formed in ookinete cultures. Cross fertilization with the male-defective Δpbs48/45 line and the female-defective Δpfs47 line further indicated that the fertilization defects of the Δpb115 lines were present in both male and female gametes. We evaluated the transmission-blocking potential of Pb115 by immunization of mice with a recombinant Pb115 fragment. In vivo mosquito feeding assay showed Pb115 immunization conferred modest, but significant transmission reducing activity with 44% reduction in infection prevalence and 39% reduction in oocyst density. Our results described functional characterization of a conserved membrane protein as a fertility factor in Plasmodium and demonstrated transmission-blocking potential of this antigen.

Artesunate and erythropoietin synergistically improve the outcome of experimental cerebral malaria.

Cerebral malaria (CM) is a severe neurological syndrome in humans and the main fatal cause of malaria. In malaria epidemic regions, despite appropriate anti-malarial treatment, 10-20% of deaths still occur during the acute phase. This is largely attributable to poor treatment access, therapeutic complexity and drug resistance; thus, developing additional clinical adjunctive therapies is an urgent necessity. In this study, we investigated the effect of artesunate (AST) and recombinant human erythropoietin (rhEPO) using an experimental cerebral malaria (ECM) model-C57BL/6 mice infected with Plasmodium berghei ANKA (PbA). Treatment with the combination of AST and rhEPO reduced endothelial activation and improved the integrity of blood brain barrier, which led to increased survival rate and reduced pathology in the ECM. In addition, this combination treatment down-regulated the Th1 response during PbA infection, which was correlated with the reduction of CCL2, TNF-α, IFN-γ, IL-12, IL-18, CXCL9 and CXCL10 levels, leading to reduced accumulation of pathogenic T cells in the brain. Meanwhile, AST and rhEPO combination led to decreased maturation and activation of splenic dendritic cells, expansion of regulatory T cells, and increased IL-10 and TGF-ß production. In conclusion, these data provide a theoretical basis for clinical adjunct therapy with rhEPO and AST in human cerebral malaria patients.

Chloroquine (CQ) exerts anti-breast cancer through modulating microenvironment and inducing apoptosis.

CQ is an anti-malaria drug, which has been used for years. However, there are published articles about its activity in anti-cancers. The aim of this approach was to look at possibility and related mechanisms of anti-breast cancer (mouse breast cancer cell line 4T1) by CQ alone. The studies of anti 4T1 in vitro and in vivo by CQ were performed. The growth of 4T1 in vitro and in vivo, survival of mice post treatment with CQ, changes of immune parameters and microenvironment in mice were evaluated. Our results demonstrate that CQ could markedly inhibit growth of 4T1 in vitro through inducing apoptosis of cells, inhibiting secretion of TGF-ß and prolong the mice survival in vivo through boosting immune system by upregulating CD8+ T cell, and through down-regulating tumor associated macrophages (TAM), myeloid derived suppressing cells (MDSC) and Tregs, in microenvironment of mice bearing tumor. This provides a new mode of action for CQ and it is therefore concluded that CQ could be with potential in breast cancer therapy.

Identification of three ookinete-specific genes and evaluation of their transmission-blocking potentials in Plasmodium berghei.

With a renewed hope for malaria elimination, interventions that prevent transmission of parasites from humans to mosquitoes have received elevated attention. Transmission-blocking vaccines (TBVs) targeting the sexual stages are well suited for this task. Here, through bioinformatic analysis, we selected two putative Plasmodium berghei ookinete-stage proteins (PBANKA_111920, and PBANKA_145770) and a previously characterized ookinete protein PBANKA_135340 (PSOP7) for evaluation of their transmission-blocking potentials. Fragments of these predicted proteins were expressed in bacteria and purified recombinant proteins were used to immunize mice. Antisera against these recombinant proteins recognized proteins of predicted sizes from ookinete lysates and localized their expression on the surface of ookinetes. Inclusion of these antisera in in vitro ookinete culture significantly inhibited ookinete formation. Mosquitoes fed on mice immunized with the recombinant proteins also showed significantly reduced oocyst densities (60.0-70.7%) and modest reductions of oocyst prevalence (10.7-37.4%). These data, together with the conservation of these genes in Plasmodium, suggest that these three ookinete proteins could be new promising targets for TBVs and are worth of future investigations in the human malaria parasites.