Optical biosensor based on weak measurement for ultra-sensitive detection of calreticulin in human serum.

A novel real-time optical phase sensing method based on the Mach-Zehnder interference principle has been proposed for the detection of calreticulin (CRT) levels in human serum samples. In this approach, anti-CRT antibodies are utilized to capture CRT molecules in serum, leading to a phase shift in both the measuring and reference arms of the system. By employing the concept of weak amplification within the framework of weak measurements, it becomes feasible to continuously monitor the response of CRT in real-time, allowing for the precise determination of serum CRT content at the picomolar level. Our achievement may pave the way in establishing CRT as a diagnostic biomarker for a wide range of medical applications, including rheumatoid arthritis.

Oxidation Analysis of l-Cysteine with a Chiral Sensor Based on Quantum Weak Measurement.

l-Cysteine, distinguished by its possession of reactive sulfhydryl groups within its molecular structure, plays a significant role in both biological systems and the pharmaceutical industry. It stands not only as a natural component integral to the constitution of glutathione but also as the principal precursor for the synthesis of l-cystine through an oxidation reaction. This study endeavors to introduce a novel approach to l-cysteine analysis, capitalizing on its optical activity, whereby an optical rotation detection system grounded in the principles of quantum weak measurement is proffered. The optical rotation angle corresponding to the concentration of chiral solutions can be accurately ascertained through spectral analysis. In practical implementation, a chiral sensing system, boasting a sensitivity of 372 nm/rad, was meticulously constructed, leveraging the concept of weak value amplification. Then, the real-time monitoring of chemical reactions involving l-cysteine and dimethyl sulfoxide was performed. Under the specific experimental conditions outlined in this investigation, it was observed that the oxidation process culminated within approximately 12 h. The application of weak measurement-based chiral sensors holds immense potential, providing robust technical support for real-time monitoring in fields such as chiral analysis and the synthesis of chiral pharmaceutical compounds.

Gender context modulation on the self-enhancement effect of vocal attractiveness evaluation.

People evaluated their own voices as sounding more attractive than others rated their voices (i.e., self-enhancement effect from the perspective of the rater, termed "SE_rater"), and people also rated their own voices as more attractive than the voices of others (i.e., self-enhancement effect from the perspective of the voice, termed "SE_voice"). The aim of the present study is to explore whether the gender context (i.e., same-sex and opposite-sex rating context) could influence the SE effect of voice attractiveness evaluation. Male and female participants were asked to rate the attractiveness of their own voices and other participants' voices, either in a same-sex session or an opposite-sex session. The results demonstrated both the SE_rater and SE_voice effect in the same-sex and opposite-sex contexts, for both male and female. More importantly, we found that the SE_rater for the male voices was significantly greater than that for the female voices in the same-sex context whereas no such difference was found in the opposite-sex context. In addition, the SE_voice effect in men was larger in the same-sex context than that in the opposite-sex context whereas the SE_voice in women was smaller in the same-sex context than that in the opposite-sex context. These findings indicated that the self-enhancement effect of vocal attractiveness was modulated by the gender context.

Mechanism underlying the self-enhancement effect of voice attractiveness evaluation: self-positivity bias and familiarity effect.

The aim of the present study was to determine whether the self-enhancement effect of voice attractiveness evaluation is due to general self-positivity bias and/or the familiarity effect. The participants were asked to rate the attractiveness of their own voice, a friend's voice and strangers' voices. In addition, a self-reference valence (SR-valence) task was adopted in the experiment. Significant self-enhancement effects in voice attractiveness ratings were demonstrated, regardless of whether the participants recognized their self-voice or not. However, the friend-enhancement effect was found in only those participants who successfully recognized their friend's voice. Moreover, a significant correlation was found between self-positivity bias in the SR-valence task and the self-enhancement effect (but not the friend-enhancement effect). Our findings suggest that both the familiarity effect and self-positivity bias account for the vocal self-enhancement effect, and the influence of self-positivity bias could be implicit. The present study thus provides empirical evidence to clarify the potential explanations for the self-enhancement of voice attractiveness assessment.

Sustained suppression of IL-18 by employing a vaccine ameliorates intestinal inflammation in TNBS-induced murine colitis.

AIM: To develop IL-18 peptide-based virus-like particle vaccines that elicit autoantibodies against IL-18 and to evaluate the in vivo effects of the vaccines in murine colitis. METHODS: Recombinant IL-18 vaccines were constructed, and the effects of the vaccines were evaluated in trinitrobenzene sulfonic acid-induced acute and chronic colitis in mice. RESULTS: Two murine IL-18 peptide-based vaccines (A and D) were developed, which induced relative long-lasting specific antibodies against IL-18. Vaccine-immunized mouse antisera could partially block IL-18-induced IFN-γ production in vitro. Mice receiving vaccine D, not vaccine A, had a significant decrease in intestinal inflammation, collagen deposition and pro-inflammatory cytokine levels in colon tissue. CONCLUSION: IL-18 vaccine may provide a potential therapeutic approach in the treatment of Crohn's disease.

Active immunization against transforming growth factor beta1 prevents hepatic fibrosis in a rat model of liver disease.

Transforming growth factor beta1 (TGF-ß1) plays an important role in hepatic fibrogenesis. In this study, we documented the effects of active immunization against TGF-ß1 on hepatic fibrosis in an animal model of chronic liver disease. BALB/c mice were immunized against 3 different peptides of TGF-ß1 ligated into hepatitis B virus core protein (HBVc). Titers of TGF-ß1 antibodies were documented by enzyme linked immunoassays and antibody activity by cell membrane receptor binding and proliferation assays. The most immunogenic recombinant HBVc + TGF-ß1 peptide (HBVc + C) then served as a vaccine in Sprague-Dawley rats with dimethylnitrosamine-induced chronic liver disease. Hepatic fibrosis was documented by serum hyaluronic acid levels, liver histology, and reverse transcriptase polymerase chain reaction for hepatic collagen I (α1) and smooth muscle alpha actin mRNA expression. Relative to control rats vaccinated with HBVc alone, recombinant HBVc + C vaccinated animals had significantly lower serum hyaluronic acid levels, less histologic evidence of hepatic fibrosis, and reduced expression of collagen I (α1) and smooth muscle alpha actin mRNA in the liver. The results of this proof-of-concept study suggest that active immunization against TGF-ß1 is a worthwhile strategy to pursue in efforts to prevent hepatic fibrosis associated with chronic liver disease.

rAed a 4: A New 67-kDa Aedes aegypti Mosquito Salivary Allergen for the Diagnosis of Mosquito Allergy.

BACKGROUND: Accurate diagnosis of mosquito allergy has been hampered by the laborious task of obtaining mosquito salivary allergens. We have previously studied 3 recombinant (r) Aedes aegypti mosquito salivary allergens: rAed a 1, rAed a 2 and rAed a 3. Here, we report the expression, purification, identification and evaluation of rAed a 4, a 67-kDa α-glucosidase. METHODS: rAed a 4 was expressed using a baculovirus/insect cell system, purified by a combination of anion- and cation-exchange chromatography, and identified by immunoblotting. A. aegypti saliva extract was prepared in our laboratory. An indirect enzyme-linked immunosorbent assay (ELISA) was developed to measure rAed a 4-specific immunoglobulin E (IgE) and IgG antibodies in sera from 13 individuals with a positive mosquito-bite test from a laboratory-reared mosquito. Sera from 18 individuals with a negative bite test served as controls. RESULTS: Purified rAed a 4 bound to the IgE in mosquito-allergic sera, as detected by ELISA and immunoblotting. The binding of rAed a 4 to IgE could be inhibited in a dose-dependent manner by the addition of an A. aegypti extract. Mosquito-allergic individuals had significantly higher mean levels of rAed a 4-specific IgE and IgG than controls. Using the mean of the controls ± 2 SD as a cut-off level, 46% of the 13 allergic individuals had a positive IgE, while none of the controls was positive (p < 0.001). CONCLUSIONS: Aed a 4 is a major allergen in mosquito saliva. Its recombinant form has the hydrolase function and can be used for the diagnosis of mosquito allergy.

Immunization against TGF-ß1 reduces collagen deposition but increases sustained inflammation in a murine asthma model.

Transforming growth factor (TGF)-ß1 is involved in the processes of airway inflammation and remodeling; however, its reported roles in asthma pathogenesis are controversial. We sought both to investigate the effects of active immunization targeting TGF-ß1 on allergen-induced airway inflammatory responses and to evaluate its possible application for asthma treatment. BALB/c mice were immunized with a virus-like-particle (VLP) vaccine presenting a TGF-ß1 peptide. For the preventive intervention of acute allergic airway inflammation, immunization was conducted before sensitization and challenges with ovalbumin (OVA), and for the therapeutic treatment of chronic inflammatory responses, immunization was initiated after inflammatory responses were established. Preventive immunization with VLPs led to increased proinflammatory IL-4, IL-13, and IL-33 levels in the bronchoalveolar lavage fluids (BALF) with no significant effects on lung tissue inflammation and airway goblet cell hyperplasia. Therapeutic treatment showed that at 24 h after the fourth 2-day challenge with OVA following 2 intraperitoneal sensitizations, airway subepithelial collagen deposition was significantly ameliorated in vaccinated mice, whereas the lung histology and cytokine profile in the BALF were not changed. In contrast, after a 4-week recovery from the last OVA challenge, the vaccinated mice's collagen deposition remained reduced, but they sustained lung-tissue inflammation and goblet-cell hyperplasia; elevated IL-13, TNF, and IFN-γ levels in the BALF; and increased airway resistance, tissue resistance, and tissue elastance. In a conclusion, the role of TGF-ß1 is complicated in allergic airway inflammatory responses. It is important to make a careful assessment in accordance with specific disease conditions when targeting TGF-ß1 for a therapeutic purpose.

Functional Myeloid-Derived Suppressor Cell Subsets Recover Rapidly after Allogeneic Hematopoietic Stem/Progenitor Cell Transplantation.

Myeloid-derived suppressor cells (MDSCs) are regulatory cell populations that have the ability to suppress effector T cell responses and promote the development of regulatory T cells (Tregs). They are a heterogeneous population of immature myeloid progenitors that include monocytic and granulocytic subsets. We postulated that given the rapid expansion of myeloid cells post-transplant, these members of the innate immune system may be important contributors to the early immune environment post-transplant. To evaluate the kinetics of recovery and function of MDSCs after allogeneic hematopoietic stem cell transplant (HSCT), 26 patients undergoing allogeneic HSCT were studied at 6 time points in the first 3 months after HSCT. Both MDSC subsets recovered between 2 and 4 weeks, well before the recovery of T and B lymphocytes. MDSC subset recovery positively correlated with T, B, and/or double-negative T cell numbers after HSCT. MDSCs isolated from patients post-transplant were functional in that they suppressed third-party CD4(+) T cell proliferation and Th1 differentiation and promoted Treg development. In conclusion, functional MDSC are present early after HSCT and likely contribute to the regulatory cell population post-transplant.

The potential protective role of caveolin-1 in intestinal inflammation in TNBS-induced murine colitis.

BACKGROUND: Caveolin-1 (Cav-1) is a multifunctional scaffolding protein serving as a platform for the cell's signal-transduction and playing an important role in inflammation. However, its role in inflammatory bowel disease is not clear. A recent study showed that Cav-1 is increased and mediates angiogenesis in dextran sodium sulphate-induced colitis, which are contradictory to our pilot findings in 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis. In the present study, we further clarified the role of Cav-1 in TNBS-induced colitis. METHODS: In BALB/c mice, acute colitis was induced by intra-rectal administration of one dose TNBS, while chronic colitis was induced by administration of TNBS once a week for 7 weeks. To assess the effects of complete loss of Cav-1, Cav-1 knockout (Cav-1-/-) and control wild-type C57 mice received one TNBS administration. Body weight and clinical scores were monitored. Colon Cav-1 and pro-inflammatory cytokine levels were quantified through ELISAs. Inflammation was evaluated through histological analysis. RESULTS: Colon Cav-1 levels were significantly decreased in TNBS-induced colitis mice when compared to normal mice and also inversely correlated with colon inflammation scores and proinflammatory cytokine levels (IL-17, IFN-γ and TNF) significantly. Furthermore, after administration of TNBS, Cav-1-/- mice showed significantly increased clinical and colon inflammatory scores and body weight loss when compared with control mice. CONCLUSIONS AND SIGNIFICANCE: Cav-1 may play a protective role in the development of TNBS-induced colitis. Our findings raise an important issue in the evaluation of specific molecules in animal models that different models may exhibit opposite results because of the different mechanisms involved.

Virus-like particles presenting interleukin-33 molecules: immunization characteristics and potentials of blockingIL-33/ST2 pathway in allergic airway inflammation.

We sought to develop an IL-33 vaccine and evaluate its efficacy in a mouse model of asthma. The full-length molecules of putative mature IL-33 were inserted into the immunodominant epitope region of hepatitis B core antigen using gene recombination techniques. The expressed chimeric protein presented as virus-like particles (VLPs) under observation using an electron microscopy. To investigate immunization characteristics of the VLPs, mice were immunized by using different doses, adjuvants, and routes. The VLPs induced sustained and high titers of IL-33-specific IgG and IgA even without the use of a conventional adjuvant, and the lowered ratio of IgG1/IgG2a in vaccinated mice indicated a shift from Th2 to Th1-like responses. To assess the vaccine effects on blocking the signaling of IL-33/ST2 pathway, mice receiving 3 vaccinations subjected to intraperitoneal sensitization and intranasal challenge with ovalbumin (OVA). Control animals received carrier or PBS in place of the vaccine. Immunization with the VLPs significantly suppressed inflammatory cell number and IL-33 level in BALF. OVA -induced goblet cell hyperplasia and lung tissue inflammatory cell infiltration were significantly suppressed in vaccinated mice. Our data indicate that IL-33 molecule-based vaccine, which may block IL-33/ST2 signaling pathway on a persistent basis, holds potential for treatment of asthma and, by extension, other diseases where overexpressed IL-33 plays a pivotal role in pathogenesis.

Employing an IL-23 p19 vaccine to block IL-23 ameliorates chronic murine colitis.

BACKGROUND: Overexpression of IL-23 has been implicated in the pathogenesis of Crohn's disease. Using vaccines to block overexpressed endogenous cytokines has emerged as a new therapeutic strategy for the long-term treatment of the disease. AIM: We sought to develop peptide-based vaccines specific to IL-23 and evaluate their effects in colitis mice. MATERIALS & METHODS: The vaccine was developed by inserting a peptide derived from mouse IL-23 p19 into the carrier protein, hepatitis B core antigen, using molecular engineering methods. One vaccine against IL-23 p19 was obtained that induced high-titered and long-lasting antibodies to IL-23 without the use of adjuvants. The inhibitory effect of vaccine-immunized serum was subsequently evaluated in vitro. To evaluate the in vivo effects, mice were subcutaneously injected with the vaccine, carrier or saline three times. Two weeks after the last injection, chronic colitis was induced in mice by seven weekly administrations with 2,4,6-trinitrobenzene sulfonic acid. RESULTS: In vitro studies revealed that serum IL-23 p19-specific IgG significantly suppressed IL-23-induced IL-17 production by splenocytes. In vivo evaluation of the effect of the vaccine in mice with chronic colitis indicated that vaccine-immunized mice exhibited a decrease in colon inflammation, collagen deposition and levels of IL-23 and IL-12 cytokines, compared with control groups. CONCLUSION: IL-23 p19 vaccine is capable of downregulating inflammatory responses in chronic murine colitis, providing a novel therapeutic approach in Crohn's disease.

The role and potential therapeutic application of myeloid-derived suppressor cells in TNBS-induced colitis.

MDSCs, a heterogeneous population of cells that expand during many pathogenic conditions, have remarkable abilities to suppress T cell responses. Their role in murine colitis, induced by TNBS and therapeutic application, remains unclear. Murine colitis was induced through intrarectally administrating TNBS, twice. MDSCs in spleen and colonic LPMCs were identified using flow cytometric analysis. In adoptive transfer, MDSCs were isolated from spleen after TNBS challenges by using microbeads or generated in vitro by coculturing bone marrow cells with HSCs and then transferred into naïve mice. Two hours later, mice were then challenged with TNBS, once/week for 2 weeks. The mice were killed four days after the second TNBS delivery, and intestinal inflammation and cytokine levels and MDSC percentages were evaluated. The percentages of CD11b+Gr-1+MDSCs and subsets (CD11b+Ly6C+ and CD11b+Ly6G+MDSCs) were increased in spleen and/or colonic LPMCs in colitis mice and also correlated with the severity of intestinal inflammation. MDSCs isolated from colitis mice suppressed the proliferation of splenocytes in vitro. Adoptive transfer of MDSCs, isolated from colitis mice or generated in vitro, decreased intestinal inflammation, levels of IFN-γ, IL-17, and TNF, and percentages of spleen MDSCs when compared with controls. MDSCs that have inhibitory function in vitro and in vivo are increased and correlated with intestinal inflammation, suggesting that they may be used as a biomarker of disease activity and a cell-based biotherapy in IBD.

Sustained suppression of IL-13 by a vaccine attenuates airway inflammation and remodeling in mice.

We previously reported that a recombinant IL-13 peptide-based virus-like particle vaccine significantly suppressed murine acute airway allergic inflammatory responses. The impact of this strategy on the development of chronic airway inflammation and remodeling has not been investigated. We evaluated whether the vaccine-mediated sustained suppression of IL-13 attenuates features of chronic airway inflammation and remodeling in mice repeatedly challenged with allergen. BALB/c mice received two intraperitoneal sensitizing injections of ovalbumin (OVA) and alum, followed by six consecutive 2-day intranasal OVA challenges at 12-day intervals and then a 4-week recovery period. Anti-IL-13 antibodies were induced with a vaccine before (preventive experiments) or after (interventional experiments) the OVA challenge commenced. Respiratory mechanics were assessed using low-frequency forced oscillation with a small animal ventilator. Cytokine concentrations in bronchoalveolar lavage fluid (BALF) and lung histology were also assessed. In the preventive experiments, vaccination significantly suppressed IL-13 concentrations, the accumulation of inflammatory cells in BALF, lung mucus production, and collagen deposition. Furthermore, vaccination significantly attenuated OVA challenge-induced increases in airway resistance, tissue resistance, and tissue elastance, both acutely and after a 4-week recovery from allergen challenges. In the interventional experiments, vaccination decreased IL-13, TGF-ß1, and IL-12p40 concentrations in BALF, as well as mucus production and collagen deposition. Chronic inflammation and sustained airway hyperresponsiveness were not significantly reversed. The persistent suppression of IL-13 with a vaccine inhibits chronic airway inflammation and the development of several key components of airway remodeling, and this intervention is more effective at early stages than later during chronic inflammation.

An IL-17 peptide-based and virus-like particle vaccine enhances the bioactivity of IL-17 in vitro and in vivo.

AIMS: To develop an IL-17 peptide-based virus-like particle vaccine that elicits autoantibodies to IL-17 and to evaluate the effects of the vaccine in mice with experimental colitis. MATERIALS & METHODS: Recombinant IL-17 vaccines were constructed by inserting selected peptides derived from mouse IL-17 into the carrier protein, hepatitis B core antigen, using molecular engineering methods. To evaluate the in vivo effects of the vaccine, mice with 2,4,6-trinitrobenzene sulfonic acid-induced chronic colitis were injected three times with the vaccine, carrier or saline after the second delivery of 2,4,6-trinitrobenzene sulfonic acid. Colon inflammation and fibrosis were evaluated by histological examination. Serum IL-17-specific IgG and colon-tissue cytokine levels were measured by ELISA. In vitro inhibition tests of sera from vaccine-immunized mice were performed using IL-17-induced IL-6 production by NIH 3T3 cells and IL-17-induced TNF production by macrophages. RESULTS: Immunization with the vaccine without the use of adjuvants induced high-titered and long-lasting antibodies to IL-17. Unexpectedly, vaccinated mice exhibited increases in colon inflammation, collagen deposition, levels of TNF and IL-17 cytokines compared with carrier and saline groups. Furthermore, in vitro study revealed that serum IL-17-specific IgG from vaccine-immunized mice significantly enhanced IL-17-induced IL-6 production and IL-17-induced TNF production dose-dependently. CONCLUSION: The IL-17 peptide-based vaccine enhances the bioactivity of IL-17 in vitro and in vivo, providing a potential immunotherapy for treatment of diseases associated with insufficient IL-17 production, such as hyper-IgE syndrome.

Targeting IL-12/IL-23 by employing a p40 peptide-based vaccine ameliorates TNBS-induced acute and chronic murine colitis.

Interleukin (IL)-12 and IL-23 both share the p40 subunit and are key cytokines in the pathogenesis of Crohn's disease. Previously, we have developed and identified three mouse p40 peptide-based and virus-like particle vaccines. Here, we evaluated the effects and immune mechanisms of the optimal vaccine in downregulating intestinal inflammation in murine acute and chronic colitis, induced by intrarectal administrations of trinitrobenzene sulfonic acid (TNBS). Mice were injected subcutaneously with vaccine, vaccine carrier or saline three times, and then intrarectally administered TNBS weekly for 2 wks (acute colitis) or 7 wks (chronic colitis). The severity of colitis was evaluated by body weight, histology and collagen and cytokine levels in colon tissue. Th1 and Th17 cells in mesenteric lymph nodes (MLN) were determined. Our results showed the vaccine induced high level and long-lasting specific IgG antibodies to p40, IL-12 and IL-23. After administrations of TNBS, vaccinated mice had significantly less body weight loss and a significant decrease of inflammatory scores, collagen deposition and expression of p40, IL-12, IL-23, IL-17, TNF, iNOS and Bcl-2 in colon tissues, compared with carrier and saline groups. Moreover, vaccinated mice exhibited a trend to lower percentages of Th1 cells in acute colitis and of Th17 cells in chronic colitis in MLN than in controls. In summary, administration of the vaccine induced specific antibodies to IL-12 and IL-23, which was associated with improvement of intestinal inflammation and fibrosis. This suggests that the vaccine may provide a potential approach for the long-term treatment of Crohn's disease.

Biological therapies of inflammatory bowel disease.

Inflammatory bowel disease (IBD) is characterized by increasing morbidity and, if suboptimally treated, poor prognosis. Recent evidence strongly suggests that dysfunctional immune responses play an important role in the pathogenesis of IBD. Therefore, immunologically downregulating the overactivated innate and adaptive immune responses may be a better approach to treat IBD than currently used pharmaceutical therapies. In recent years, many new biological therapies have been developed. These therapies are shown to be effective for inducing remission, preventing complications, improving life quality of the patients, and reducing hospitalization and surgical rates. This article introduces and discusses these new biological agents that have been used effectively in clinic for IBD patients.

AMPK agonist downregulates innate and adaptive immune responses in TNBS-induced murine acute and relapsing colitis.

AMP-activated protein kinase (AMPK), a cellular energy sensor, has been reported to participate in modulating inflammatory responses, but its role in intestinal inflammation remains unclear. IBD has been characterized by excessive innate and adaptive immune responses. Here, the roles of 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), an agonist of AMPK, in regulating immune responses of experimental colitis were investigated. In vitro effects of AICAR on LPS-induced macrophage activation and Th1 and Th17 differentiation, as well as in vivo effects of AICAR in mice with 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, were explored. In acute colitis, daily AICAR treatment commenced 2 days after TNBS delivery (day 1), while in relapsing colitis, AICAR treatment commenced after three weekly TNBS administrations. Colon inflammation, production of proinflammatory cytokines and NF-κB activation in colon tissues, and Th1 and Th17 cell populations in lamina propria mononuclear cells (LPMCs) and mesenteric lymph node cells (MLNs) were assayed. Results show that AICAR significantly inhibited in vitro LPS-induced macrophage activation and Th1 and Th17 cell differentiation. Administration of AICAR was therapeutically effective in ameliorating acute and relapsing experimental colitis, as shown by reduced body weight loss and significant attenuation in colon histological inflammation. Moreover, this treatment inhibited NF-κB activation in macrophages, and reduced levels of TNF, Th1- and Th17-type cytokines, and Th1 and Th17 cell populations in LPMCs and MLNs. AICAR-initiated AMPK activation may act as a central downregulator in ongoing innate and adaptive immune responses of murine colitis, providing a novel therapeutic approach in the treatment of IBD.