Contribution of parasite and host genotype to immunopathology of schistosome infections.

BACKGROUND: The role of pathogen genotype in determining disease severity and immunopathology has been studied intensively in microbial pathogens including bacteria, fungi, protozoa and viruses but is poorly understood in parasitic helminths. The medically important blood fluke Schistosoma mansoni is an excellent model system to study the impact of helminth genetic variation on immunopathology. Our laboratory has demonstrated that laboratory schistosome populations differ in sporocyst growth and cercarial production in the intermediate snail host and worm establishment and fecundity in the vertebrate host. Here, we (i) investigate the hypothesis that schistosome genotype plays a significant role in immunopathology and related parasite life history traits in the vertebrate mouse host and (ii) quantify the relative impact of parasite and host genetics on infection outcomes. METHODS: We infected BALB/c and C57BL/6 mice with four different laboratory schistosome populations from Africa and the Americas. We quantified disease progression in the vertebrate host by measuring body weight and complete blood count (CBC) with differential over a 12-week infection period. On sacrifice, we assessed parasitological (egg and worm counts, fecundity), immunopathological (organ measurements and histopathology) and immunological (CBC with differential and cytokine profiles) characteristics to determine the impact of parasite and host genetics. RESULTS: We found significant variation between parasite populations in worm numbers, fecundity, liver and intestine egg counts, liver and spleen weight, and fibrotic area but not in granuloma size. Variation in organ weight was explained by egg burden and intrinsic parasite factors independent of egg burden. We found significant variation between infected mouse lines in cytokine levels (IFN-γ, TNF-α), eosinophils, lymphocytes and monocyte counts. CONCLUSIONS: This study showed that both parasite and host genotype impact the outcome of infection. While host genotype explains most of the variation in immunological traits, parasite genotype explains most of the variation in parasitological traits, and both host and parasite genotypes impact immunopathology outcomes.

Contribution of parasite and host genotype to immunopathology of schistosome infections.

Background: The role of pathogen genotype in determining disease severity and immunopathology has been studied intensively in microbial pathogens including bacteria, fungi, protozoa, and viruses, but is poorly understood in parasitic helminths. The medically important blood fluke Schistosoma mansoni is an excellent model system to study the impact of helminth genetic variation on immunopathology. Our laboratory has demonstrated that laboratory schistosome populations differ in sporocyst growth and cercarial production in the intermediate snail host and worm establishment and fecundity in the vertebrate host. Here, we (i) investigate the hypothesis that schistosome genotype plays a significant role in immunopathology and related parasite life history traits in the vertebrate mouse host and (ii) quantify the relative impact of parasite and host genetics on infection outcomes. Methods: We infected BALB/c and C57BL/6 mice with four different laboratory schistosome populations from Africa and the Americas. We quantified disease progression in the vertebrate host by measuring body weight and complete blood count (CBC) with differential over an infection period of 12 weeks. On sacrifice, we assessed parasitological (egg and worm counts, fecundity), immunopathological (organ measurements and histopathology), and immunological (CBC with differential and cytokine profiles) characteristics to determine the impact of parasite and host genetics. Results: We found significant variation between parasite populations in worm numbers, fecundity, liver and intestine egg counts, liver and spleen weight, and fibrotic area, but not in granuloma size. Variation in organ weight was explained by egg burden and by intrinsic parasite factors independent of egg burden. We found significant variation between infected mouse lines in cytokines (IFN-γ, TNF-α), eosinophil, lymphocyte, and monocyte counts. Conclusions: This study showed that both parasite and host genotype impact the outcome of infection. While host genotype explains most of the variation in immunological traits, parasite genotype explains most of the variation in parasitological traits, and both host and parasite genotype impact immunopathology outcomes.

Contribution of parasite and host genotype to immunopathology of schistosome infections.

Background: The role of pathogen genotype in determining disease severity and immunopathology has been studied intensively in microbial pathogens including bacteria, fungi, protozoa, and viruses, but is poorly understood in parasitic helminths. The medically important blood fluke Schistosoma mansoni is an excellent model system to study the impact of helminth genetic variation on immunopathology. Our laboratory has demonstrated that laboratory schistosome populations differ in sporocyst growth and cercarial production in the intermediate snail host and worm establishment and fecundity in the vertebrate host. Here, we (i) investigate the hypothesis that schistosome genotype plays a significant role in immunopathology and related parasite life history traits in the vertebrate mouse host and (ii) quantify the relative impact of parasite and host genetics on infection outcomes. Methods: We infected BALB/c and C57BL/6 mice with four different laboratory schistosome populations from Africa and the Americas. We quantified disease progression in the vertebrate host by measuring body weight and complete blood count (CBC) with differential over an infection period of 12 weeks. On sacrifice, we assessed parasitological (egg and worm counts, fecundity), immunopathological (organ measurements and histopathology), and immunological (CBC with differential and cytokine profiles) characteristics to determine the impact of parasite and host genetics. Results: We found significant variation between parasite populations in worm numbers, fecundity, liver and intestine egg counts, liver and spleen weight, and fibrotic area, but not in granuloma size. Variation in organ weight was explained by egg burden and by intrinsic parasite factors independent of egg burden. We found significant variation between infected mouse lines in cytokines (IFN-γ, TNF-α), eosinophil, lymphocyte, and monocyte counts. Conclusions: This study showed that both parasite and host genotype impact the outcome of infection. While host genotype explains most of the variation in immunological traits, parasite genotype explains most of the variation in parasitological traits, and both host and parasite genotype impact immunopathology outcomes.

Establishment of a secondary infection laboratory model of Echinococcus shiquicus metacestode using BALB/c mice and Mongolian jirds (Meriones unguiculatus).

Echinococcus shiquicus is peculiar to the Qinghai­Tibet plateau of China. Research on this parasite has mainly focused on epidemiological surveys and life cycle studies. So far, limited laboratory studies have been reported. Here, experimental infection of E. shiquicus metacestode in BALB/c mice and Mongolian jirds (Meriones unguiculatus) was carried out to establish alternative laboratory animal models. Intraperitoneal inoculation of metacestode material containing protoscoleces (PSCs) obtained from infected plateau pikas were conducted on BALB/c mice. Furthermore, metacestode material without PSCs deriving from infected BALB/c mice was intraperitoneally inoculated to Mongolian jirds. Experimental animals were dissected for macroscopic and histopathological examination. The growth of cysts in BALB/c mice was infiltrative, and they invaded the murine entire body. Most of the metacestode cysts were multicystic, but a few were unilocular. The cysts contained sterile vesicles, which had no PSCs. The metacestode materials were able to successfully infect new mice. In the jirds model, E. shiquicus cysts were typically formed freely in the peritoneal cavity; the majority of these cysts were free while a small portion adhered loosely to nearby organs. The proportion of fertile cysts was high, and contained many PSCs. The PSCs produced in Mongolian jirds also successfully infected new ones, which confirms that jirds can serve as an alternative experimental intermediate host. In conclusion, a laboratory animal infection was successfully established for E. shiquicus using BALB/c mice and Mongolian jirds. These results provide new models for the in-depth study of Echinococcus metacestode survival strategy, host interactions and immune escape mechanism.

The effect of the anti-leukemia inhibitory factor on the immune system in the Balb/c mice bearing breast cancer induced with 4T1 cells.

BACKGROUND: Breast cancer is one of the most common cancers. Leukemia inhibitory factor (LIF) is considered as one of the effective factors in the growth of breast cancer, and anti-leukemia inhibitory factor antibody is considered as one of the treatment options for this type of cancer. METHODS: Mice models of breast cancer were made with 4T1 cell line and were randomly divided into four groups. The first group included the mice that received anti-LIF (Anti LIF group). The mice in the second group received anti-LIF and doxorubicin (Anti LIF & DOX). The mice in the third group received only doxorubicin (DOX). Finally, the mice in the fourth group did not receive any intervention. 22 days after tumor induction, some of the mice were killed, and their tumor tissues, lymph nodes, and spleens were separated for evaluating P53, Caspase-3, TIM-3, LAG-3, CTLA-4, and PD-1 genes expression. The percentage of regulatory T cells and level of interferon gamma (IFN-γ) and transforming growth factor-beta (TGF-ß) were evaluated. The rest of the mice were kept to check the tumor size and their survival rate. RESULTS: The proposed intervention did not have any significant effect on the tumor growth and the survival rate. However, the expression of P53 gene and Caspase-3 in the tumor tissue of the Anti LIF group had a significant enhancement. In tumor tissues and lymph nodes, the expression of T-bet, PD-1, TIM-3, and LAG-3 genes in the Anti LIF group showed a significant increase. There was no significant difference between groups in the percentage of regulatory T cells and level of IFN-γ and TGF-ß. CONCLUSIONS: The proposed interventions were able to have a direct effect on tumors, but no significant effect was observed on the immune system.

The Effect of Potassium Nitrate Supplementation on the Force and Properties of Extensor digitorum longus (EDL) Muscles in Mice.

Adding potassium nitrate (KNO3) to the diet improves the physiological properties of mammalian muscles (rebuilds weakened muscle, improves structure and functionality). The aim of this study was to investigate the effect of KNO3 supplementation in a mouse model. BALB/c mice were fed a KNO3 diet for three weeks, followed by a normal diet without nitrates. After the feeding period, the Extensor digitorum longus (EDL) muscle was evaluated ex vivo for contraction force and fatigue. To evaluate the possible pathological changes, the histology of EDL tissues was performed in control and KNO3-fed groups after 21 days. The histological analysis showed an absence of negative effects in EDL muscles. We also analyzed 15 biochemical blood parameters. After 21 days of KNO3 supplementation, the EDL mass was, on average, 13% larger in the experimental group compared to the controls (p < 0.05). The muscle-specific force increased by 38% in comparison with the control group (p < 0.05). The results indicate that KNO3 has effects in an experimental mouse model, showing nitrate-diet-induced muscle strength. This study contributes to a better understanding of the molecular changes in muscles following nutritional intervention and may help develop strategies and products designated to treat muscle-related issues.

The BALB/c Mouse Model for the Evaluation of Therapies to Treat Infections with Aerosolized Burkholderia pseudomallei.

Burkholderia pseudomallei, the causative agent of the disease melioidosis, has been isolated from the environment in 45 countries. The treatment of melioidosis is complex, requiring lengthy antibiotic regimens, which can result in the relapse of the disease following treatment cessation. It is important that novel therapies to treat infections with B. pseudomallei be assessed in appropriate animal models, and discussions regarding the different protocols used between laboratories are critical. A 'deep dive' was held in October 2020 focusing on the use of the BALB/c mouse model and the inhalational route of infection to evaluate new antibiotic therapies.

A blood-brain barrier-penetrating AAV2 mutant created by a brain microvasculature endothelial cell-targeted AAV2 variant.

Upon systemic administration, adeno-associated virus serotype 9 (AAV9) and the capsid variant PHP.eB show distinct tropism for the central nervous system (CNS), whereas AAV2 and the capsid variant BR1 transduce brain microvascular endothelial cells (BMVECs) with little transcytosis. Here, we show that a single amino acid substitution (from Q to N) in the BR1 capsid at position 587 (designated BR1N) confers a significantly higher blood-brain barrier (BBB) penetration capacity to BR1. Intravenously infused BR1N showed significantly higher CNS tropism than BR1 and AAV9. BR1 and BR1N likely use the same receptor for entry into BMVECs; however, the single amino acid substitution has profound consequences on tropism. This suggests that receptor binding alone does not determine the final outcome in vivo and that further improvements of capsids within predetermined receptor usage are feasible.

A preliminary study on placental damage associated to experimental neosporosis in BALB/c mice.

Neospora caninum is a protozoan parasite which can infect a range of animals, including dogs, cattle, and sheep. Bovine neosporosis, which mainly causes abortion in cattle, results in substantial economic losses worldwide. To study the effects of N. caninum infection on the placenta, a pregnant mouse model for N. caninum infection was established. The litter size (8.6 ± 1.5) and the number of live pups (6.4 ± 1.8) of infected dams were significantly lower compared with those of non-infected dams. Trophoblast cell shrinkage and a large number of apoptosomes were detected in the placentas of the infected group. The parasite load in the placental tissue was significantly higher with time after infection. Likewise, apoptosis of placental trophoblast cells significantly increased with time after infection. Among the 66 apoptotic genes detected in this study, eight genes, including Bcl-2, were significantly differentially expressed by about > tenfold in infected and uninfected mice. The expression of BAX and tumor necrosis factor-alpha (TNF-α) was upregulated in the placental cells of the infected mice, whereas the expression of BCL-2 was downregulated. Enzyme-linked immunosorbent assays (ELISAs) showed that apoptotic protease caspase-3 level was significantly increased in placental cell suspension, and insulin-like growth factor (IGF)-2 level was significantly reduced. Acetylcholine (ACH) and placental prolactin (PL) levels were initially decreased but eventually increased. In summary, infection of mice with N. caninum caused apoptotic damage to the placental tissues, cells, and genes and affected the normal physiological functions of placenta, which may largely explain the adverse pregnancy outcomes caused by N. caninum infection in mice.

From Mouse to Man: N-Methyl-d-Aspartic Acid Receptor Activation as a Promising Pharmacotherapeutic Strategy for Autism Spectrum Disorders.

The BALB/c mouse displays hypersensitivity to behavioral effects of MK-801 (dizocilpine), a noncompetitive N-methyl-d-aspartic acid (NMDA) receptor "open-channel" blocker, and shows both no preference for an enclosed stimulus mouse over an inanimate object and reduced social interaction with a freely behaving stimulus mouse. NMDA receptor agonist interventions improved measures of social preference and social interaction of the BALB/c mouse model of autism spectrum disorder (ASD). A "proof of principle/proof of concept" translational 10-week clinical trial with 8-week of active medication administration was conducted comparing 20 DSM-IV-TR-diagnosed older adolescent/young adult patients with ASD randomized to once-weekly pulsed administration (50 mg/d) versus daily administration of d-cycloserine (50 mg/d). The results showed that d-cycloserine, a partial glycine agonist, was well tolerated, the 2 dosing strategies did not differ, and improvement was noted on the "lethargy/social withdrawal" and "stereotypic behavior" subscales of the Aberrant Behavior Checklist. NMDA receptor activation contributes to the regulation of mTOR signaling, a pathologic point of convergence in several monogenic syndromic forms of ASD. Furthermore, both NMDA receptor hypofunction and imbalance between NMDA receptor activation mediated by GluN2B and GluN2A-containing NMDA receptors occur as "downstream" consequences of several genetically unrelated abnormalities associated with ASD. NMDA receptor-subtype selective "positive allosteric modulators (PAMs)" are particularly appealing medication candidates for future translational trials.

Ultrasound-assisted glycation and the allergenicity of α-lactalbumin.

BACKGROUND: Ultrasound-assisted glycation is a promising method for decreasing the allergenicity of α-lactalbumin (ALA). However, there is a lack of in vivo studies on the allergenicity of ultrasound-assisted glycated ALA. In this study, the effects of the ultrasound-assisted glycation of ALA on the allergenicity and intestinal microflora were characterized using a BALB/c mouse model. RESULTS: Increased immunoglobulin -G/ immunoglobulin-E (IgG/IgE) and interleukin-4/6 (IL-4/6) secretions, and reduced interferon-γ (IFN-γ) secretions were found in the serum of ALA sensitized and challenged, mice in comparison with a control group. However, there was no significant difference between the mice fed with ultrasound-assisted glycated ALA and the control group. Mice that were sensitized and challenged with ALA showed disrupted intestinal microflora, manifesting in significantly decreased Firmicutes and significantly increased Proteobacteria. It was found that 100ALA-gal could maintain the intestinal microflora of mice in a normal state. Pearson's rank correlation showed that Proteobacteria and Spirochaetota were correlated positively with the IL-4/IL-6 level and were correlated negatively with the expression of IFN-γ. Proteobacteria were also significantly positively correlated with IL-6 and negatively correlated with IFN-γ (P < 0.05). CONCLUSION: These results suggested that ultrasound-assisted glycation on ALA can maintain the intestinal microflora in a normal state thus balancing the proportion of Th1/Th2 to decrease allergic reaction. © 2022 Society of Chemical Industry.

Neospora caninum surface antigen 1 is a major determinant of the pathogenesis of neosporosis in nonpregnant and pregnant mice.

Introduction: NcSAG1 is one of most widely investigated antigens of Neospora caninum in various research fields. Such studies demonstrated the proficiency of NcSAG1 in the regulatory process of parasite adhesion and invasion of host cells. Accordingly, the contribution of NcSAG1 to the pathogenesis of neosporosis can undoubtedly be extrapolated, but direct evidence is lacking. Herein, we provide the first successful attempt at the gene disruption of NcSAG1 and novel data on the invasion and virulence potentials of N. caninum in vitro and in vivo. Methods: The disruption of the NcSAG1 gene was applied using the CRISPR/Cas9 system and confirmed by PCR, western blot and indirect fluorescent antibody tests as NcSAG1 knockout parasites (NcSAG1KO). Then, we investigated the role of NcSAG1 in the growth kinetics of the parasite in vitro. Results and discussion: The deletion of the NcSAG1 gene significantly decreased the infection rate and reduced the egress rate of the parasite. An in vivo study using nonpregnant female and male BALB/c mice revealed a significantly higher survival rate and lower body weight change in the group infected with the NcSAG1KO parasite than in the parental strain (Nc-1)-infected group. Regarding the vertical transmission model of BALB/c mice, the absence of the NcSAG1 gene significantly enhanced the survival of pups and greatly lowered the parasite burden in the brains of pups. In conclusion, our study suggested NcSAG1 as a key molecule in the pathogenesis of N. caninum.

A replication-competent smallpox vaccine LC16m8Δ-based COVID-19 vaccine.

Viral vectors are a potent vaccine platform for inducing humoral and T-cell immune responses. Among the various viral vectors, replication-competent ones are less commonly used for coronavirus disease 2019 (COVID-19) vaccine development compared with replication-deficient ones. Here, we show the availability of a smallpox vaccine LC16m8Δ (m8Δ) as a replication-competent viral vector for a COVID-19 vaccine. M8Δ is a genetically stable variant of the licensed and highly effective Japanese smallpox vaccine LC16m8. Here, we generated two m8Δ recombinants: one harbouring a gene cassette encoding the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) glycoprotein, named m8Δ-SARS2(P7.5-S)-HA; and one encoding the S protein with a highly polybasic motif at the S1/S2 cleavage site, named m8Δ-SARS2(P7.5-SHN)-HA. M8Δ-SARS2(P7.5-S)-HA induced S-specific antibodies in mice that persisted for at least six weeks after a homologous boost immunization. All eight analysed serum samples displayed neutralizing activity against an S-pseudotyped virus at a level similar to that of serum samples from patients with COVID-19, and more than half (5/8) also had neutralizing activity against the Delta/B.1.617.2 variant of concern. Importantly, most serum samples also neutralized the infectious SARS-CoV-2 Wuhan and Delta/B.1.617.2 strains. In contrast, immunization with m8Δ-SARS2(P7.5-SHN)-HA elicited significantly lower antibody titres, and the induced antibodies had less neutralizing activity. Regarding T-cell immunity, both m8Δ recombinants elicited S-specific multifunctional CD8+ and CD4+ T-cell responses even after just a primary immunization. Thus, m8Δ provides an alternative method for developing a novel COVID-19 vaccine.

BET-Independent Murine Leukemia Virus Integration Is Retargeted In Vivo and Selects Distinct Genomic Elements for Lymphomagenesis.

Moloney murine leukemia virus (MLV) infects BALB/c mice and induces T-cell lymphoma in mice. Retroviral integration is mediated by the interaction of the MLV integrase (IN) with members of the bromodomain and extraterminal motif (BET) protein family (BRD2, BRD3, and BRD4). The introduction of the W390A mutation into MLV IN abolishes the BET interaction. Here, we compared the replication of W390A MLV to that of wild-type (WT) MLV in adult BALB/c mice to study the role of BET proteins in replication, integration, and tumorigenesis in vivo. Comparing WT and W390A MLV infections revealed similar viral loads in the blood, thymus, and spleen cells. Interestingly, W390A MLV integration was retargeted away from GC-enriched genomic regions. However, both WT MLV- and W390A MLV-infected mice developed T-cell lymphoma after similar latencies represented by an enlarged thymus and spleen and multiorgan tumor infiltration. Integration site sequencing from splenic tumor cells revealed clonal expansion in all WT MLV- and W390A MLV-infected mice. However, the integration profiles of W390A MLV and WT MLV differed significantly. Integrations were enriched in enhancers and promoters, but compared to the WT, W390A MLV integrated less frequently into enhancers and more frequently into oncogene bodies such as Notch1 and Ppp1r16b. We conclude that host factors direct MLV in vivo integration site selection. Although BET proteins target WT MLV integration preferentially toward enhancers and promoters, insertional lymphomagenesis can occur independently from BET, likely due to the intrinsically strong enhancer/promoter of the MLV long terminal repeat (LTR). IMPORTANCE In this study, we have shown that the in vivo replication of murine leukemia virus happens independently of BET proteins, which are key host determinants involved in retroviral integration site selection. This finding opens a new research line in the discovery of alternative viral or host factors that may complement the dominant host factor. In addition, our results show that BET-independent murine leukemia virus uncouples insertional mutagenesis from gene enhancers, although lymphomagenesis still occurs despite the lack of an interaction with BET proteins. Our findings also have implications for the engineering of BET-independent MLV-based vectors for gene therapy, which may not be a safe alternative.

Optimized Extraction of cAMP From Jujube by Ultra-High Pressure Technology and the Anti-allergic Effect for Peanut Allergy Mouse.

Jujube contains abundant cyclic adenosine monophosphate (cAMP). In contrast, the extraction technology of cAMP from jujube is still to be explored. In this study, the ultra-high pressure extraction (UHPE) conditions for obtaining the maximum cAMP yield from jujube were optimized. Orthogonal array design (OAD) was applied to evaluate the effects of three variables (pressure, pressure-holding time, and liquid-to-solid ratio) by UHPE on cAMP yield. The results showed that the optimal cAMP yield (1223.2 µg/g) was derived at 300 MPa, 20 min duration, and a liquid-to-solid ratio of 2.5 ml/g. In addition, as an important functional ingredient in jujube, cAMP has potential anti-allergic effect. To develop the functional characteristics of jujube, the effect of cAMP was characterized in vivo with the Balb/c mouse model of peanut allergy, which was established by subcutaneous injection of crude peanut protein extract (PN). The results showed that treatment with cAMP in PN-sensitized mice suppressed the lesions in jejunal tissues and allergic symptoms and restored spleen index. Meanwhile, cAMP treatment reduced serum levels of specific immunoglobulin E (IgE), histamine, as well as interleukin-4 (IL-4) and stimulated the secretion of tumor necrosis factor-α (TNF-α), whereas the serum levels of interleukin-10 (IL-10) were not affected. Our results suggested that cAMP has an anti-allergic effect in PN-sensitized mice.

Thorium promotes lung, liver and kidney damage in BALB/c mouse via alterations in antioxidant systems.

Thorium (232Th), long lived (14.05 billion years) most stable thorium isotope, is thrice naturally abundant than uranium. 232Th occurs as rocky deposits and black monazite sands on the earth's crust geographically distributed in coastal South India and other places globally. Monazite sand comprises of cerium and large quantities of radioactive thorium. The environmental hazard lies in monazite rich area being termed as High Background Radiation Area (HBRA). In this study, we mimicked the HBRA under controlled chamber conditions using thorium oxalate as a thorium source for BALB/c mice exposure. Furthermore, sequential radio-disintegration of 232 Th leads to thoron (220Rn), the noble gas and other daughter products/progeny predominantly via alpha decay/emissions. Such progeny tend to attach to aerosol and dust particles having potential inhalation hazard followed by alpha emissions and damages that we evaluated in mouse lung tissues post thoron inhalation. Secondly, along with the radio disintegration and alpha emission, high energy gamma is also generated that can travel to various distant organs through the systemic circulation, as significant findings of our study as damages to the liver and kidney. The mechanistic findings include the damages to the hematological, immunological and cellular antioxidant systems along with activation of canonical NF-κß pathway via double stranded DNA damage.

Comparative characterization of microRNAs of Schistosoma japonicum from SCID mice and BALB/c mice: Clues to the regulation of parasite growth and development.

Schistosomiasis, caused by a parasite with a wide range of mammalian hosts, remains one of the most prevailing parasitic diseases in the world. While numerous studies have reported that the growth and reproduction of schistosomes in immunodeficient mice was significantly retarded, the underlying molecular mechanisms have yet to be revealed. In this study, we comparatively analyzed the microRNA expression of Schistosoma japonicum derived from SCID and BALB/c mice on the 35th day post-infection by high-throughput RNA sequencing as prominent morphological abnormalities had been observed in schistosomes from SCID mice when compared with those from BALB/c mice. The results revealed that more than 72% and 61% of clean reads in the small RNA libraries of female and male schistosomes, respectively, could be mapped to the selected miRs in the miRBase or the sequences of species-specific genomes. Further analysis identified 122 miRNAs using TPM >0.01 as the threshold value, including 75 known and 47 novel miRNAs, 96 of which were commonly expressed across all the four tested schistosome libraries. Comparative analysis of the libraries of schistosomes from SCID and BALB/c mice identified 15 differentially expressed miRNAs (5 up-regulated and 10 down-regulated) among females and 16 among males (9 up-regulated and 7 down-regulated). Integrated analysis of the two sets of differentially expressed miRNAs of female and male worms identified 2 miRNAs (sja-miR-3488 and sja-miR-novel_29) that overlapped between female and male datasets. Prediction of miRNA targets and Gene Ontology (GO) term enrichment analysis of the predicted target genes revealed that these genes were involved in some important biological processes, such as nucleic acid metabolic process, macromolecule modification, and cellular aromatic compound metabolic process. The predicted target genes were further matched to the differentially expressed genes in male and female schistosomes from the above two hosts, obtaining 7 genes that may be responsible for regulating the growth, development and sex maturation of schistosomes. Taken together, this study provides the first identification of differentially expressed miRNAs in schistosomes from SCID and BALB/c mice. These miRNAs and their predicted target mRNAs are probably involved in the regulation of development, growth, and maturation of schistosomes. Therefore, this study expands our understanding of schistosome development regulation and host-parasite relationship, and also provides a valuable set of potential anti-schistosomal targets for prevention and control of schistosomiasis.

Evaluation of allergenicity of cow milk treated with enzymatic hydrolysis through a mouse model of allergy.

Cow milk (CM) allergy is a worldwide concern. Currently, few studies have been performed on the immunoreactivity of CM and fewer still on the antigenicity of CM in vivo and in vitro. In this study, we assessed the potential allergenicity of enzymatically hydrolyzed CM using in vitro ELISA and oral sensitization and challenge of BALB/c mice. Alcalase-, Protamex-, and Flavourzyme-treated CM (all from Novozymes) diminished IgE binding capacity, with greatest reductions of 56.31%, 50.62%, and 56.45%, respectively. Allergic symptoms and levels of total IgG1 were reduced, and allergic inflammation of the lung, jejunum, and spleen was relieved. Moreover, the numbers of CD8+ T and B220+ cells decreased, and the balance of CD4+ T/CD8+ T cells was effectively regulated. These findings suggest that the potential allergenicity of CM was reduced by enzymatic hydrolysis, and our research will lay a solid foundation for developing high-quality hypoallergenic CM products.

Fascin Regulates the Hippo Pathway and Is Important for Melanoma Development.

BACKGROUND/AIM: Fascin, an actin-bundling protein, plays an essential role in cancer metastasis. The Hippo pathway is critical for carcinogenesis and cancer stem cell self-renewal. Mammalian STE20-like kinase (MST) is a core component of the Hippo pathway. However, whether fascin and MST2 affect melanoma remain largely unknown. This study aimed to investigate the role of fascin and MST2 in melanoma development. MATERIALS AND METHODS: Surgically excised skin melanomas and the adjacent non-tumorous skin tissue from 30 cases were analyzed using immunohistochemistry for fascin and MST2. The melanoma cell line WM793 was employed for fascin and MST2 knock-down followed by western blotting, and melanoma xenografting in BALB/c mice. RESULTS: Immunohistochemistry revealed increased expression of fascin and decreased expression of MST2 in melanoma. The reverse correlation of fascin and MST2 was statistically significant. Fascin siRNA upregulated MST2 expression; however, MST2 siRNA did not significantly affect fascin expression in the WM793. WM793 xenografting followed by fascin knock-down inhibited tumor growth significantly in the animal study. CONCLUSION: Fascin is a regulator of the Hippo pathway and plays an important role in melanoma development. Therefore, fascin could be a potential therapeutic target for melanoma.

Sphingomyelin liposome bearing whole Leishmania lysate antigens induce strong Th2 immune response in BALB/c mice.

OBJECTIVES: Whole Leishmania lysate antigens (WLL) has been shown to be effective to tackle leishmaniasis in murine models. Although liposomes can be considered as promising vaccines, the activity of phospholipase-A (PLA) in WLL, breeds difficulties to preparing stable liposomal WLL. One strategy to overcome this shortcoming is to use lipids such as sphingomyelin (SM) which is resistant against PLA. This study aim is formulating stable SM liposomes containing WLL and comparing their adjuvant effects with another first generation vaccine , i.e. solube Leishmania Antigen (SLA) liposomes in BALB/c mice. MATERIALS AND METHODS: BALB/c mice were immunized subcutaneously, three times with 2-week intervals, with Empty-liposome (E-lipo), Particulate WLL, Liposome-WLL, Liposome-SLA and control Buffer, three times every 2-week. Protection was assessed through measuring the swollen footpads and the load of parasites in the spleen. Other factors were used to assess the response of immune system by means of IgG subclasses, IL-4 and IFN-γ levels and intracellular cytokine assay in cultured splenocytes. RESULTS: Although liposomal WLL were stable in terms of physicochemical properties, mice received Liposome-WLL did not reduce footpad swelling. The load of parasites in spleen and levels of IL-4- were also higher compared to other immunized groups. In terms of IgG isotypes, no considerable difference observed in mice received Liposome-WLL or other formulations. CONCLUSION: Liposome-WLL could be a suitable vaccine delivery system when a Th2 response is desired. Also, further studies are warranted to fully understand the role of sphingomyelin in inducing an immune response.