Screening of Multiple Myeloma by Polyclonal Rabbit Anti-Human Plasmacytoma Cell Immunoglobulin

Antibody-based immunotherapy has been effectively used for tumor treatment. However, to date, only a few tumor-associated antigens (TAAs) or therapeutic targets have been identified. Identification of more immunogenic antigens is essential for improvements in multiple myeloma (MM) diagnosis and therapy. In this study, we synthesized a polyclonal antibody (PAb) by immunizing rabbits with whole human plasmacytoma ARH-77 cells and identified MM-associated antigens, including enlonase, adipophilin, and HSP90s, among others, via proteomic technologies. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay showed that 200 µg/mL PAb inhibits the proliferation of ARH-77 cells by over 50% within 48 h. Flow cytometric assay indicated that PAb treatment significantly increases the number of apoptotic cells compared with other treatments (52.1% vs. NS, 7.3% or control rabbit IgG, 9.9%). In vivo, PAb delayed tumor growth and prolonged the lifespan of mice. Terminal deoxynucleotidyl transferase dUTP nick end labeling assay showed that PAb also induces statistically significant changes in apoptosis compared with other treatments (P<0.05). We therefore conclude that PAb could be used for the effective screening and identification of TAA. PAb may have certain anti-tumor functions in vitro and in vivo. As such, its combination with proteomic technologies could be a promising approach for sieving TAA for the diagnosis and therapy of MM.     

A Homogeneous,High-Throughput Assay for Phosphatidylinositol 5-Phosphate 4-Kinase with a Novel,Rapid Substrate Preparation

Phosphoinositide kinases regulate diverse cellular functions and are important targets for therapeutic development for diseases, such as diabetes and cancer. Preparation of the lipid substrate is crucial for the development of a robust and miniaturizable lipid kinase assay. Enzymatic assays for phosphoinositide kinases often use lipid substrates prepared from lyophilized lipid preparations by sonication, which result in variability in the liposome size from preparation to preparation. Herein, we report a homogeneous 1536-well luciferase-coupled bioluminescence assay for PI5P4Kα. The substrate preparation is novel and allows the rapid production of a DMSO-containing substrate solution without the need for lengthy liposome preparation protocols, thus enabling the scale-up of this traditionally difficult type of assay. The Z’-factor value was greater than 0.7 for the PI5P4Kα assay, indicating its suitability for high-throughput screening applications. Tyrphostin AG-82 had been identified as an inhibitor of PI5P4Kα by assessing the degree of phospho transfer of γ-³²P-ATP to PI5P; its inhibitory activity against PI5P4Kα was confirmed in the present miniaturized assay. From a pilot screen of a library of bioactive compounds, another tyrphostin, I-OMe tyrphostin AG-538 (I-OMe-AG-538), was identified as an ATP-competitive inhibitor of PI5P4Kα with an IC₅₀ of 1 µM, affirming the suitability of the assay for inhibitor discovery campaigns. This homogeneous assay may apply to other lipid kinases and should help in the identification of leads for this class of enzymes by enabling high-throughput screening efforts.     

Increased Expression of IL-22 Is Associated with Disease Activity in Behcet’s Disease

Objective

Interleukin (IL)-22 has been reported to be involved in the development of autoimmune diseases. This study aimed to analyze the expression and potential role of IL-22 in the pathogenesis of Behcet’s disease (BD).

Methods

The levels of IL-22 in patient sera or supernatants of cultured peripheral blood mononuclear cells (PBMCs) and CD4⁺T cells were detected by enzyme-linked immunosorbent assay (ELISA). Flow cytometry was used to evaluate the frequency of IL-22–producing CD4⁺ T cells. IL-22 mRNA from erythema nodosum skin lesions was examined using real time quantitative RT-PCR.

Results

BD patients with active uveitis showed a significantly higher expression of IL-22 in the supernatants of stimulated PBMCs and CD4⁺T cells compared with BD patients without active uveitis and normal controls. An increased frequency of IL-22-producing CD4⁺T cells was also found in BD patients with active uveitis. IL-22 mRNA expression was elevated in erythema nodosum skin lesions. In BD patients, a high IL-22 level in the supernatant of stimulated PBMCs correlated with the presence of retinal vasculitis and erythema nodosum.

Conclusions

IL-22 was associated with disease activity in BD and correlated with the presence of small vessel inflammation, suggesting that it may be involved in its pathogenesis.     

Enhancing Specific-Antibody Production to the ragB Vaccine with GITRL That Expand Tfh,IFN-γ+ T Cells and Attenuates Porphyromonas gingivalis Infection in Mice

The outer membrane protein RagB is one of the major virulence factors of the periodontal pathogen Porphyromonas gingivalis (P. gingivalis). In order to induce protective immune response against P. gingivalis infection, an mGITRL gene-linked ragB DNA vaccine (pIRES-ragB-mGITRL ) was constructed. Six-week-old female BALB/c mice were immunized with pIRES-ragB-mGITRL through intramuscular injection and then challenged by subcutaneous injection in the abdomen with P. gingivalis. RagB-specific antibody-forming cells were evaluated by an Enzyme-linked immunosorbent spot, and specific antibody was determined by enzyme-linked immunosorbent assay. In addition, the frequencies of Tfh and IFN-γ⁺ T cells in spleen were measured using flow cytometer, and the levels of IL-21 and IFN-γ mRNA or proteins were detected by real time RT-PCR or ELISA. The data showed that the mGITRL-linked ragB DNA vaccine induced higher levels of RagB-specific IgG in serum and RagB-specific antibody-forming cells in spleen. The frequencies of Tfh and IFN-γ⁺ T cells were obviously expanded in mice immunized by pIRES-ragB-mGITRL compared with other groups (pIRES or pIRES-ragB ). The levels of Tfh and IFN-γ⁺ T cells associated cytokines were also significantly increased in pIRES-ragB-mGITRL group. Therefore, the mice immunized with ragB plus mGITRL showed the stronger resistant to P. gingivalis infection and a significant reduction of the lesion size caused by P. gingivalis infection comparing with other groups. Taken together, our findings demonstrated that intramuscular injection of DNA vaccine ragB together with mGITRL induced protective immune response dramatically by increasing Tfh and IFN-γ⁺ T cells and antibody production to P. gingivalis.     

CD106 Identifies a Subpopulation of Mesenchymal Stem Cells with Unique Immunomodulatory Properties

Mesenchymal stem cells (MSCs) reside in almost all of the body tissues, where they undergo self-renewal and multi-lineage differentiation. MSCs derived from different tissues share many similarities but also show some differences in term of biological properties. We aim to search for significant differences among various sources of MSCs and to explore their implications in physiopathology and clinical translation. We compared the phenotype and biological properties among different MSCs isolated from human term placental chorionic villi (CV), umbilical cord (UC), adult bone marrow (BM) and adipose (AD). We found that CD106 (VCAM-1) was expressed highest on the CV-MSCs, moderately on BM-MSCs, lightly on UC-MSCs and absent on AD-MSCs. CV-MSCs also showed unique immune-associated gene expression and immunomodulation. We thus separated CD106⁺cells and CD106⁻cells from CV-MSCs and compared their biological activities. Both two subpopulations were capable of osteogenic and adipogenic differentiation while CD106⁺CV-MSCs were more effective to modulate T helper subsets but possessed decreased colony formation capacity. In addition, CD106⁺CV-MSCs expressed more cytokines than CD106⁻CV-MSCs. These data demonstrate that CD106 identifies a subpopulation of CV-MSCs with unique immunoregulatory activity and reveal a previously unrecognized mechanism underlying immunomodulation of MSCs.     

STK39 Polymorphism Is Associated with Essential Hypertension: A Systematic Review and Meta-Analysis

Background

A recent genome-wide association study identified STK39as a candidate gene for blood pressure (BP) in Europeans. Subsequently, several studies have attempted to replicate the association across different ethnic populations. However, the results have been inconsistent.

Objective and Methods

We performed a meta-analysis to elucidate the association between the STK39 rs3754777 polymorphism (or proxy) and hypertension. Published literature from PubMed and Embase databases were retrieved and pooled odds ratio (OR) with 95% confidence interval (CI) was calculated using fixed- or random-effects model.

Results

Using appropriate inclusion/exclusion criteria, we identified 10 studies that included 21, 863 hypertensive cases and 24, 480 controls from different ethnicities. The meta-analysis showed a significant association of STK39 rs3754777 variant with hypertension (OR = 1.10, 95%CI = 1.06–1.15, p = 7.95×10⁻⁶). Further subgroup analysis by ethnicity suggested that the association was significant in Europeans (OR = 1.08, 95% CI = 1.03–1.14, p = 0.002) and in East Asians (OR = 1.16, 95%CI = 1.07–1.25, p = 4.34×10⁻⁴), but not in Africans (OR = 1.01, 95%CI 0.80–1.27, p = 0.932). We further confirmed the positive association by sensitivity analysis. No publication bias was detected (Begg’s test, p = 0.721; Egger’s test, p = 0.744).

Conclusions

The present meta-analysis confirms the significant association of STK39 polymorphism with susceptibility to hypertension in Europeans and East Asians. Future studies should include gene–gene and gene–environment interactions to investigate the identified association.     

Prevalence,Awareness, Medication,Control, and Risk Factors Associated with Hypertension in Bai Ethnic Group in Rural China: The Yunnan Minority Eye Study

Purpose

This study aimed to assess the prevalence, awareness, treatment, and control of hypertension and their associated factors among Bai ethnic population in the rural China.

Methods

A population-based survey was conducted in 2010 with a randomly cluster sampling in rural communities in Dali, southwest China. A total of 2133 adults aged 50 or above were interviewed, and their blood pressure, height, weight and waist circumference were measured. Hypertension was defined as a mean SBP≥140 mmHg and/or DBP≥90 mmHg, and/or current use of antihypertensive medications.

Results

The prevalence of hypertension was 42.1% (899/2133), and the age- and gender-adjusted prevalence was 40.0%. Among the hypertensive participants, 28.4% (255/899)were aware of their condition, while 24.6% (221/899) took antihypertensive medications, with only 7.5% (67/899) of those achieving blood pressure control (<140/90 mmHg). Risk factors for hypertension were older age, smoking, alcohol drinking, family history of HBP, overweight, and obesity, while protective factors included being lean, and having finished senior high school or above.

Conclusions

Hypertension prevalence is high among the population of Bai ethnic group in China, while the associated risk factors of hypertension include overweight/obesity, cigarette smoking, history of hypertension, and older age. The percentages of hypertensive participants aware of their hypertension and those taking antihypertensive medications were low with an incredibly low proportion of hypertensive patients who kept their hypertension under control. It is suggested that health education and hypertension screening programs be carried out in the area for the high blood pressure prevention and control.     

Educational Level,Obesity and Incidence of Diabetes among Chinese Adult Men and Women Aged 18–59 Years Old: An 11-Year Follow-Up Study

Objective

To determine whether educational level and overweight/obesity was associated with the development of diabetes among Chinese adult men and women.

Methods

A cohort (2000–2011) of 10 704 participants aged 18–59 years (8 238 men, 2 466 women) in Qingdao Port Health Study (QPHS) were recruited in this study. The personal lifestyle, height, weight, waist circumference, resting heart rate, blood pressure, fasting blood glucose, total cholesterol, triglycerides and plasma uric acid were collected annually in a comprehensive health checkup program. Cox proportional hazards regression models were used to estimate the association of factors and incidence of diabetes.

Results

During 110 825 person-years of follow-up, 1 056 new onset cases (9.5 per 1 000 person-years) of diabetes were identified. With normal weight as reference, the multiple-adjusted hazard ratio (HR) (95%CI) of diabetes was 1.69(1.38–2.09) for overweight and 2.24(1.66–3.02) for obesity among men, which was 1.81(1.12–2.92) and 2.58(1.37–4.86) among women, respectively. Compared with the participants with high educational level, those with low educational level had a higher risk of diabetes (multiple-adjusted HR (95%CI): 1.43(1.11–1.86)) among men. The association was not found among women and the adjusted HR (95%CI) of diabetes was 1.56(0.89–2.76). The increased risks of low educational level were independent of mediators among men, through normal weight (P for trend = 0.0313) and overweight (P for trend = 0.0212) group but not obesity group (P for trend = 0.0957).

Conclusion

Baseline overweight/obesity was an independent risk factor for diabetes for both men and women. Low educational level was adversely associated with incidence of diabetes through normal weight, overweight and obesity groups, with the association being substantially attenuated by mediating factors only in the obesity group among men. The association was not found among women.     

Molecular and Clinical Characteristics of Clonal Complex 59 Methicillin-Resistant Staphylococcus aureus Infections in Mainland China

Detailed molecular analyses of Clonal Complex 59 (CC59) methicillin-resistant Staphylococcus aureus (MRSA) isolates from children in seven major cities across Mainland China were examined. A total of 110 CC59 isolates from invasive and non-invasive diseases were analyzed by multilocus sequence typing (MLST), Staphylococcus cassette chromosome mec (SCCmec) typing, staphylococcal protein A (spa) typing and pulsed-field gel electrophoresis (PFGE). Antibiotics susceptibilities, carriage of plasmids and 42 virulence genes and the expression of virulence factors were examined. ST59 (101/110, 91.8%) was the predominant sequence type (ST), while single locus variants (SLVs) belonging to ST338 (8/110, 7.3%) and ST375 (1/110, 0.9%) were obtained. Three SCCmec types were found, namely type III (2.7%), type IV (74.5%) and type V (22.7%). Seven spa types including t437, which accounted for 87.3%, were determined. Thirteen PFGE types were obtained. PFGE types A and B were the major types totally accounting for 81.8%. The dominant clone was ST59-t437-IVa (65.5%), followed by ST59-t437-V (14.5%). The positive rate of luks-PV and lukF-PV PVL encoding (pvl) gene was 55.5%. Plasmids were detected in 83.6% (92/110) of the strains. The plasmid size ranging from 23.4 kb to 50 kb was most prevalent which accounted for 83.7% (77/92). A significantly lower expression of hla was found in ST59-t437-IVa compared with ST59-t437-V. Among the 110 cases, 61.8% of the patients were less than 1 year old. A total of 90 cases (81.8%) were community-associated (CA) infections whereas 20 cases (18.2%) were hospital-associated (HA) infections. Out of the 110 patients, 36.4% (40/110) were diagnosed with invasive infectious diseases in which ST59-t437-IVa accounted for 67.5% (27/40). In brief, ST59-t437-IVa was proved as the dominant clone in CC59 MRSA strains. The carriage rate of pvl gene was high. CC59 MRSA could result in CA and HA infections. The majortiy of MRSA infection children were in young age.     

Total HIV/AIDS Expenditures in Dehong Prefecture,Yunnan Province in 2010: The First Systematic Evaluation of Both Health and Non-Health Related HIV/AIDS Expenditures in China

Objective

We assessed HIV/AIDS expenditures in Dehong Prefecture, Yunnan Province, one of the highest prevalence regions in China, and describe funding sources and spending for different categories of HIV-related interventions and at-risk populations.

Methods

2010 HIV/AIDS expenditures in Dehong Prefecture were evaluated based on UNAIDS’ National AIDS Spending Assessment methodology.

Results

Nearly 93% of total expenditures for HIV/AIDS was contributed by public sources. Of total expenditures, 52.7% was allocated to treatment and care, 24.5% to program management and administration and 19.8% to prevention. Spending on treatment and care was primarily allocated to the treatment of opportunistic infections. Most (40.4%) prevention spending was concentrated on most-at-risk populations, injection drug users (IDUs), sex workers, and men who have sex with men (MSM), with 5.5% allocated to voluntary counseling and testing. Prevention funding allocated for MSM, partners of people living with HIV and prisoners and other confined populations was low compared to the disproportionate burden of HIV/AIDS in these populations. Overall, people living with HIV accounted for 57.57% of total expenditures, while most-at-risk populations accounted for only 7.99%.

Conclusions

Our study demonstrated the applicability of NASA for tracking and assessing HIV expenditure in the context of China, it proved to be a useful tool in understanding national HIV/AIDS response from financial aspect, and to assess the extent to which HIV expenditure matches epidemic patterns. Limited funding for primary prevention and prevention for MSM, prisoners and partners of people living with HIV, signal that resource allocation to these key areas must be strengthened. Comprehensive analyses of regional and national funding strategies are needed to inform more equitable, effective and cost-effective HIV/AIDS resource allocation.