Interleukin-32γ suppresses allergic airway inflammation in mouse models of asthma.

Asthma is a chronic airway inflammatory disease typically associated with T helper cell type 2 (Th2) cytokines. IL-32, first reported as an inducer of tumor necrosis factor (TNF)-α, is an inflammatory cytokine involved in various autoinflammatory diseases, viral infection, and cancer-related inflammation. However, the role of IL-32γ in asthma has not been clearly elucidated. In this study, the levels of IL-32γ in sputum from patients with asthma were measured by ELISA, and IL-32γ function was investigated in murine models of asthma with human IL-32γ-overexpressed transgenic (IL-32γ TG) mice. The therapeutic effect of recombinant IL-32γ (rIL-32γ) on allergic inflammation was also evaluated through bronchoalveolar lavage fluid analysis and histopathologic examinations. Sputum IL-32γ levels from patients with asthma were lower than those from healthy control subjects. In an acute mouse model of asthma, IL-32γ TG mice exhibited significantly reduced airway inflammation compared with that in wild-type mice. The production of Th1 cytokines, such as TNF-α and IFN-γ, and Th2 cytokines, such as IL-4, IL-5, and IL-13, was decreased in the lungs of IL-32γTG mice. On the contrary, the expression of IL-10 and IL-10-producing CD11b(+) monocytic cells was significantly increased in the lungs of ovalbumin-sensitized IL-32γ TG mice. In addition, rIL-32γ treatment revealed a suppressive effect on the airway inflammation in a chronic mouse model of asthma. The results of this study suggest that IL-32γ may have a preventive role in the development of allergic airway inflammation and could be a potential novel therapeutic target for bronchial asthma.

IL-32γ overexpression accelerates streptozotocin (STZ)-induced type 1 diabetes.

Interleukin-32 (IL-32) is a cytokine produced by T lymphocytes, natural killer (NK) cells, monocytes and epithelial cells. There are five splicing variants (α, ß, γ, δ, and ε) and IL-32γ is the most active isoform. We generated human IL-32γ transgenic (IL-32γ TG) mice, displaying a high level of IL-32γ expression in the pancreas. We investigated the effect of IL-32γ on streptozotocin (STZ)-induced type 1 diabetes model using IL-32γ TG mice. After a suboptimal diabetogenic dose of STZ administration, IL-32γ TG mice showed significantly increased blood glucose level comparing with that of wild type (WT) mice at day 5. Inflammatory cytokines levels such as, IL-6, TNFα, IFNγ and IL-1ß, in pancreas and liver lysates were accessed by a specific cytokine ELISA. The proinflammatory cytokines were significantly enhanced in the pancreas of IL-32γ TG mice comparing to that of WT mice whereas those cytokines levels in liver of IL-32γ TG and WT mice were not changed by STZ. These data indicate that the overexpression of IL-32γ contributes to initial islet ß-cells injury and inflammation in pancreas and aggravates STZ-induced type 1 diabetes.

Contradictory functions (activation/termination) of neutrophil proteinase 3 enzyme (PR3) in interleukin-33 biological activity.

IL-1 family ligand does not possess a typical hydrophobic signal peptide and needs a processing enzyme for maturation. The maturation process of IL-33 (IL-1F11), a new member of the IL-1 family ligand, remains unclear. Precursor IL-33 ligand affinity column isolates neutrophil proteinase 3 (PR3) from human urinary proteins. PR3 is a known IL-1 family ligand-processing enzyme for IL-1ß (IL-1F2) and IL-18 (IL-1F4), including other inflammatory cytokines. We investigated PR3 in the maturation process of precursor IL-33 because we isolated urinary PR3 by using the precursor IL-33 ligand affinity column. PR3 converted inactive human and mouse precursor IL-33 proteins to biological active forms; however, the increase of PR3 incubation time abrogated IL-33 activities. Unlike caspase-1-cleaved precursor IL-18, PR3 cut precursor IL-33 and IL-18 at various sites and yielded multibands. The increased incubation period of PR3 abated mature IL-33 in a time-dependent manner. The result is consistent with the decreased bioactivity of IL-33 along with the increased PR3 incubation time. Six different human and mouse recombinant IL-33 proteins were expressed by the predicted consensus amino acid sequence of PR3 cleavage sites and tested for bioactivities. The human IL-33/p1 was highly active, but human IL-33/p2 and p3 proteins were inactive. Our results suggest the dual functions (activation/termination) of PR3 in IL-33 biological activity.

Effect of recombinant α1-antitrypsin Fc-fused (AAT-Fc)protein on the inhibition of inflammatory cytokine production and streptozotocin-induced diabetes.

α1-Antitrypsin (AAT) is a member of the serine proteinase inhibitor family that impedes the enzymatic activity of serine proteinases, including human neutrophil elastase, cathepsin G and neutrophil proteinase 3. Here, we expressed recombinant AAT by fusing the intact AAT gene to the constant region of IgG1 to generate soluble recombinant AAT-Fc protein. The recombinant AAT-Fc protein was produced in Chinese hamster ovary (CHO) cells and purified using mini-protein A affinity chromatography. Recombinant AAT-Fc protein was tested for antiinflammatory function and AAT-Fc sufficiently suppressed tumor necrosis factor (TNF)-α-induced interleukin (IL)-6 in human peripheral blood mononuclear cells (PBMCs) and inhibited cytokine-induced TNFα by different cytokines in mouse macrophage Raw 264.7 cells. However, AAT-Fc failed to suppress lipopolysaccharide-induced cytokine production in both PBMCs and macrophages. In addition, our data showed that AAT-Fc blocks the development of hyperglycemia in a streptozotocin-induced mouse model of diabetes. Interestingly, we also found that plasma-derived AAT specifically inhibited the enzymatic activity of elastase but that AAT-Fc had no inhibitory effect on elastase activity.

Paradoxical effects of constitutive human IL-32{gamma} in transgenic mice during experimental colitis.

Inflammatory cytokines mediate inflammatory bowel diseases (IBDs) and cytokine blocking therapies often ameliorate the disease severity. IL-32 affects inflammation by increasing the production of IL-1, TNFα, and several chemokines. Here, we investigated the role of IL-32 in intestinal inflammation by generating a transgenic (TG) mouse expressing human IL-32γ (IL-32γ TG). Although IL-32γ TG mice are healthy, constitutive serum and colonic tissue levels of TNFα are elevated. Compared with wild-type (WT) mice, IL-32γ TG mice exhibited a modestly exacerbated acute inflammation early following the initiation of dextran sodium sulfate (DSS)-induced colitis. However, after 6 d, there was less colonic inflammation, reduced tissue loss, and improved survival rate compared with WT mice. Associated with attenuated tissue damage, colonic levels of TNFα and IL-6 were significantly reduced in the IL-32γ TG mice whereas IL-10 was elevated. Cultured colon explants from IL-32γ TG mice secreted higher levels of IL-10 compared with WT mice and lower levels of TNFα and IL-6. Constitutive levels of IL-32γ itself in colonic tissues were significantly lower following DSS colitis. Although the highest level of serum IL-32γ occurred on day 3 of colitis, IL-32 was below constitutive levels on day 9. The ability of IL-32γ to increase constitutive IL-10 likely reduces TNFα, IL-6, and IL-32 itself accounting for less inflammation. In humans with ulcerative colitis (UC), serum IL-32 is elevated and colonic biopsies contain IL-32 in inflamed tissues but not in uninvolved tissues. Thus IL-32γ emerges as an example of how innate inflammation worsens as well as protects intestinal integrity.

Engineering of a human kringle domain into agonistic and antagonistic binding proteins functioning in vitro and in vivo.

Here, we report the development of target-specific binding proteins based on the kringle domain (KD) ( approximately 80 residues), a ubiquitous modular structural unit occurring across eukaryotic species. By exploiting the highly conserved backbone folding by core residues, but using extensive sequence variations in the seven loop regions of naturally occurring human KDs, we generated a synthetic KD library on the yeast cell surface by randomizing 45 residues in the loops of a human KD template. We isolated KD variants that specifically bind to anticancer target proteins, such as human death receptor 4 (DR4) and/or DR5, and that function as agonists to induce apoptotic cell death in several cancer cell lines in vitro and inhibit tumor progression in mouse models. Combined treatments with KD variants possessing different recognition sites on the same target protein exerted synergisitic tumoricidal activities, compared to treatment with individual variants. In addition to the agonists, we isolated an antagonistic KD variant that binds human tumor necrosis factor-alpha (TNFalpha) and efficiently neutralizes TNFalpha-induced cytotoxicity in vitro and in vivo. The KD scaffold with seven flexible loops protruding from the central core was strongly sequence-tolerant to mutations in the loop regions, offering a potential advantage of distinct binding sites for target recognition on the single domain. Our results suggest that the KD scaffold can be used to develop target-specific binding proteins that function as agonists or antagonists toward given target molecules, indicative of their potential use as biotherapeutics.

A Robust Quadruple Protein-Based Indirect ELISA for Detection of Antibodies to African Swine Fever Virus in Pigs.

African swine fever (ASF) emerged in domestic pigs and wild boars in China in 2018 and rapidly spread to neighboring Asian countries. Currently, no effective vaccine or diagnostic tests are available to prevent its spread. We developed a robust quadruple recombinant-protein-based indirect enzyme-linked immunosorbent assay (QrP-iELISA) using four antigenic proteins (CD2v, CAP80, p54, and p22) to detect ASF virus (ASFV) antibodies and compared it with a commercial kit (IDvet) using ASFV-positive and -negative serum samples. The maximum positive/negative value was 24.033 at a single antigen concentration of 0.25 µg/mL and quadruple ASFV antigen combination of 1 µg/mL at a 1:100 serum dilution. Among 70 ASFV-positive samples, 65, 67, 65, 70, 70, and 14 were positive above the cut-offs of 0.121, 0.121, 0.183, 0.065, 0.201, and 0.122, for CD2v, CAP80, p54, p22-iELISA, QrP-iELISA, and IDvet, respectively, with sensitivities of 92.9%, 95.7%, 92.9%, 100%, 100%, and 20%, respectively, all with 100% specificity. The antibody responses in QrP-iELISA and IDvet were similar in pigs infected with ASFV I. QrP-iELISA was more sensitive than IDvet for early antibody detection in pigs infected with ASFV II. These data provide a foundation for developing advanced ASF antibody detection kits critical for ASF surveillance and control.

Identification of constitutively active interleukin 33 (IL-33) splice variant.

IL-33/IL-1F11 is a new member of the IL-1 family ligand and provokes T helper-type immune responses. IL-33 is the ligand of ST2 and IL-1 receptor accessory protein (IL-1RAcP) that triggers nuclear factor-κ light chain enhancer of activated B cells (NF-κB) and MAPK signaling. We discovered a novel short splice variant of IL-33 that was termed spIL-33. The new spIL-33 lacks exon 3 containing a proposed caspase-1 cleavage site. We isolated spIL-33 cDNA from the Huh7 human hepatocarcinoma cell line and expressed the recombinant spIL-33 protein in Escherichia coli. The recombinant spIL-33 and pro-IL-33 were not cleaved by caspase-1, unlike IL-18 (IL-1F4). The recombinant spIL-33 was constitutively active, and spIL-33-induced inflammatory cytokine production was caspase-1-independent in HMC-1 and Raw 264.7 cells. The recombinant spIL-33 induced the phosphorylation of IL-1 receptor-associated kinase (IRAK1), NF-κB, p38 MAPK, p44/42 MAPK, and JNK in a time- and dose-dependent manner. Anti-ST2 monoclonal antibody specifically blocked the spIL-33-induced cytokine production. In this study, we identified and characterized a new IL-33 splice variant, which was a constitutively active IL-33 isoform. The existence of constitutively active spIL-33 suggests that the biological activity of IL-33 could be triggered by diverse stimulations during immune responses. Further investigation of the spIL-33 expression pattern may contribute to understanding the involvement of IL-33 in inflammatory disorders.

Elevated interleukin-32 expression in granulomatosis with polyangiitis.

OBJECTIVE: To evaluate the role of IL-32 in granulomatosis with polyangiitis (GPA) patients and the relationship between IL-32 and disease activity, as PR3 has the ability to bind and activate IL-32, which has been described as a novel cytokine that induces inflammatory cytokines. METHODS: We investigated the level of IL-32, PR3, TNF-α and IL-6 in GPA patients by using ELISA. Northern blot was used to analyse the level of IL-32 mRNA in leucocytes of GPA patients. The intracellular colocalization of IL-32 and PR3 in leucocytes was examined by IF staining. RESULTS: We observed that IL-32 and PR3 levels in GPA patients were increased significantly when compared with normal individuals and each was tightly associated (P < 0.001). Northern blot analysis revealed that the mRNA level of IL-32 was prominently elevated in leucocytes of GPA patients. The intracellular colocalization of IL-32 and PR3 in leucocytes from GPA patients vs normal individuals was verified by IF staining. CONCLUSION: IL-32 level was elevated in GPA patients but its level was changed by treatment response. IL-32 could be an index in GPA and play a role in the aetiology of GPA.

Characterizing antiviral mechanism of interleukin-32 and a circulating soluble isoform in viral infection.

Interleukin-32 (IL-32) is an inflammatory cytokine, and its activity is associated with various auto-inflammatory disorders as well as infectious pathogens such as Mycobacterium tuberculosis, and viral infections. However, the precise antiviral mechanism of IL-32 remains unclear. We assessed the IL-32 level in the sera of H1N1 influenza A patients and IL-32 level was significantly elevated. Next we examined the antiviral activity of recombinant IL-32γ (rIL-32γ) with WISH cells infected by vesicular stomatitis virus (VSV) but no antiviral activity was observed. Therefore we investigated the supernatant of rIL-32-treated THP-1 cells since this cell line effectively responded to rIL-32γ. The supernatant of rIL-32-treated THP-1 cell possessed an antiviral effect and in addition, an agonistic monoclonal antibody further enhanced a specific antiviral activity of rIL-32γ. The fractionation and mass spectrometer analysis of the THP-1 cell supernatant revealed that the antiviral activity of rIL-32γ is via a THP-1 cell-produced factor, transferrin, rather than the direct effects of rIL-32γ on epithelial cells. We also characterized a secreted soluble IL-32γ protein in serum of IL-32γ transgenic mouse (TG), but not in that of IL-32α TG. The present results suggest that IL-32γ expression and its genetic variation in individual could be an important aspect of viral infections.

Clinical characteristics and the expression profiles of inflammatory cytokines/cytokine regulatory factors in asymptomatic patients with nodular gastritis.

BACKGROUND: Nodular gastritis (NG) has been reported in adult dyspeptic patients, whereas information on NG in asymptomatic patients is limited. AIMS: To evaluate the prevalence, clinico-epidemiological characteristics, and expression profiles of inflammatory cytokines or cytokine regulatory factors of NG in asymptomatic adults. METHODS: A cross-sectional study was conducted prospectively using 2,579 consecutive asymptomatic subjects who underwent screening esophagogastroduodenoscopy. The expression of inflammatory cytokines or cytokine regulatory factors in the gastric mucosa of NG patients was evaluated using immunofluorescence staining. RESULTS: NG was diagnosed in 52 patients (2.0%) and showed a predilection for females (M:F = 1:1.89) and young adults (median age: 34 years; range: 25-51 years). All NG patients were positive for Helicobacter pylori infection. Based on multivariate analysis, the risk of NG was increased in patients younger than 40 years (OR, 7.57; 95% CI, 3.76-15.24) and of the female gender (OR, 2.12; 95% CI; 1.05-4.28). Immunofluorescent staining for interleukin (IL)-1ß, IL-10, IL-18, IL-18 binding protein, IL-32, IL-33, and neutrophil proteinase 3 (PR3) was performed on cryosections of gastric mucosa. Interestingly, the expression of PR3 was highly increased in the gastric biopsies from asymptomatic NG patients but was expressed infrequently in the controls. CONCLUSIONS: Asymptomatic NG is associated with H. pylori infection, and a predilection for this condition exists in young females. The PR3 expression of gastric mucosa might play an important role in the pathogenesis of NG.

Neutrophil proteinase 3 induces diabetes in a mouse model of glucose tolerance.

Type 1 diabetes is considered to be an autoimmune disease in which T cells attack pancreatic islet cells. Impaired glucose tolerance with type 2 diabetes has been classified as an obesity-associated metabolic syndrome. However, recent studies have revealed that type 2 diabetes is an autoinflammatory disease due to an imbalance of inflammatory cytokine production and related molecular components that cause inflammation. Insulin-like growth factor (IGF) and the insulin-like growth factor-binding protein-3 (IGFBP3) system are known to be involved in the development of experimental diabetic nephropathy, and urinary IGFBP3 protease activity has been observed in patients with type 2 diabetes. A serine protease was found to be responsible for the proteolytic activity in diabetic urine; however, the identity of the precise enzyme remains unknown. We investigated neutrophil proteinase 3 (PR3) to see whether it has specific enzymatic activity associated with insulin-like growth factor-1 and IGFBP3. In our study, both molecules were sufficiently degraded, which leads us to believe that PR3 may induce insulin resistance in the mouse model utilized. In addition, we found that PR3 in the urine of diabetic patients similarly affects insulin resistance. Moreover, PR3-immunized mice had an increase in glucose clearance due to inhibition of PR3 activity. As such, PR3 can be considered as an inflammatory enzyme directly linking inflammation to type 2 diabetes through downregulation of insulin-like growth factor-1/IGFBP3.

Development of an indirect ELISA against African swine fever virus using two recombinant antigens, partial p22 and p30.

African swine fever (ASF) is a highly fatal viral disease affecting pigs. It is caused by the ASF virus (ASFV), and causes serious economic losses to the swine industry worldwide, including in Korea. Commercially available enzyme-linked immunosorbent assay (ELISA) kits for detecting anti-ASFV antibodies are used for the diagnosis and surveillance of ASF. In this study, an ELISA was developed to detect anti-ASFV antibodies using two recombinant proteins, p22 and p30, from genotype II ASFV. Recombinant transmembrane domain-deleted p22 (p22∆TM) and p30 were expressed in E.coli vector system pET32a and mixed for use as antigens in indirect ELISA. The p22∆TM/p30-based indirect ELISA was validated using 31 sera from genotype I ASFV-infected pigs and 1133 sera from uninfected pigs. Area under the curve of this test was 0.999 [95 % concentration interval 0.992-1.000], and sensitivity and specificity were 93.5 % and 99.8 %, respectively. The between run coefficient of variation for internal quality control serum was 6.61 %. In the seroconversion analysis, the p22∆TM/p30-based indirect ELISA showed equal or better ability to detect antibodies in pigs experimentally challenged with ASFV p72 genotypes I and II (p < 0.05). In conclusion, the p22∆TM/p30-based indirect ELISA is a reliable diagnostic method for detecting anti-ASFV antibodies.

Evaluation of swine protection with foot-and-mouth disease O1/Campos and O/Primorsky/2014 vaccines against the O Mya-98 lineage virus from East Asia.

Two type O commercial vaccines, the O1/Campos and O/Primorsky/2014 vaccines, were studied to evaluate the in vivo efficacy in pigs against heterologous virus challenge with the O/SKR/Jincheon/2014 virus (O/SEA/Mya-98 lineage) isolated in Korea in 2014. The in vivo challenge results indicated that both vaccines induced a high heterologous virus neutralization test (VNT) titer by a single injection and successfully protected specific pathogen-free (SPF) pigs from challenge infection. To determine the optimal vaccination age, a field trial with each vaccine was conducted with three one-shot-vaccinated groups that were injected at 8, 12, or 14 weeks of age and one two-shot-vaccinated group that was injected at 8 and 12 weeks of age in the pig farms. In these field trials, the improved serological performance at 20 and 24 weeks of age expected with vaccination at 12 or 14 weeks of age was not observed, although improved serological results were expected as the result of decreasing interference of maternally derived antibodies (MDAs), as MDAs waned with age. In addition, delayed vaccination resulted in MDA depletion at 14 weeks of age. Therefore, the optimal age for primary vaccination with two different formulated vaccines was 8 weeks old in pigs, considering that MDAs could provide a protective immunity against foot-and-mouth disease (FMD) infection. Prolonged significantly higher VNT titers of immunized pigs were demonstrated in the two-shot-vaccinated groups. In total, the effectiveness of the two vaccines was demonstrated through efficacy tests and field trials in pigs.

Isolation and Genetic Characterization of African Swine Fever Virus from Domestic Pig Farms in South Korea, 2019.

On 17 September 2019, the first outbreak of African swine fever in a pig farm was confirmed in South Korea. By 9 October, 14 outbreaks of ASF in domestic pigs had been diagnosed in 4 cities/counties. We isolated viruses from all infected farms and performed genetic characterization. The phylogenetic analysis showed that all of fourteen ASFV isolates in South Korea belong to genotype II and serogroup 8. Additionally, all isolates had an intergenic region (IGR) II variant with additional tandem repeat sequences (TRSs) between the I73R and I329L genes and showed characteristics of central variable region (CVR) 1 of the B602L gene and IGR 1 of MGF 505 9R/10R genes. These are identical to the genetic characteristics of some European isolates and Chinese isolates.

The interference effect of maternally-derived antibodies on the serological performance of pigs immunized with a foot-and-mouth disease oil emulsion vaccine.

To control foot-and-mouth disease (FMD) outbreaks that originated in Jincheon County in South Korea between 2014 and 2015, several commercial vaccines were studied for their efficacy and serological performance in the field. In this study, the efficacy of the O SKR 7/10 vaccine was evaluated by challenge with the FMD virus (FMDV) O/Jincheon/SKR/2014 (O Jincheon), which has the same O/SEA/Mya-98 lineage as the O/SKR/7/10 strain that was isolated in 2010 in South Korea, in FMD-seronegative pigs. Full protection against the O Jincheon virus was demonstrated as early as 14 days postvaccination, which was explained by the strong serological relationship (r1 value: ≥ 0.92) between the O Jincheon and O SKR 2010 viruses. However, in the field trial, no satisfactory serological elevations against FMDV were observed, even in the double-vaccinated groups. Therefore, it can be concluded that the O SKR 7/10 vaccine may need to be improved to overcome the interference effects from the high levels of maternally-derived antibodies generated due to the mandatory nationwide vaccination of sows in South Korea.

Identification of the most active interleukin-32 isoform.

Cytokines are crucial in host defence against pathogens such as bacteria, viruses, fungi and parasites. A newly described cytokine, interleukin-32 (IL-32), induces various proinflammatory cytokines (tumour necrosis factor-alpha, IL-1beta, IL-6) and chemokines in both human and mouse cells through the nuclear factor-kappaB and p38 mitogen-activated protein kinase inflammatory signal pathway. The IL-32 primarily acts on monocytic cells rather than T cells. In an attempt to isolate the IL-32 soluble receptor, we used an IL-32 ligand-affinity column to purify neutrophil proteinase 3, which is a serine proteinase involved in many inflammatory diseases. IL-32 has biological activity associated with Mycobacterium tuberculosis and chronic proinflammatory diseases such as rheumatoid arthritis. IL-32 is transcribed as six alternative splice variants and the biological activity of each individual isoform remains unknown. Here, we cloned the complementary DNA of the four IL-32 isoforms (alpha, beta, gamma and delta) that are the most representative IL-32 transcripts. To produce recombinant protein with a high yield, the amino acids of two cysteine residues were mutated to serine residues, because serine residues are not conserved among different species. The multi-step purified recombinant IL-32 isoform proteins were assessed for their biological activities with different cytokine assays. The gamma isoform of IL-32 was the most active, although all isoforms were biologically active. The present study will provide a specific target to neutralize endogenous IL-32, which may contribute to basic and clinical immunology.

Differences in circulating dendritic cell subtypes in pregnant women, cord blood and healthy adult women.

Different subtypes of dendritic cells (DC) influence the differentiation of naíve T lymphocytes into T helper type 1 (Th1) and Th2 effector cells. We evaluated the percentages of DC subtypes in peripheral blood from pregnant women (maternal blood) and their cord blood compared to the peripheral blood of healthy non pregnant women (control). Circulating DC were identified by flow cytometry as lineage (CD3, CD14, CD16, CD19, CD20, and CD56)-negative and HLA-DR-positive cells. Subtypes of DC were further characterized as myeloid DC (CD11c(+)/CD123(+/-)), lymphoid DC (CD11c(-)/CD123(+++)) and less differentiated DC (CD11c(-)/CD123(+/-)). The frequency of DC out of all nucleated cells was significantly lower in maternal blood than in control (P<0.001). The ratio of myeloid DC/lymphoid DC was significantly higher in maternal blood than in control (P<0.01). HLA-DR expressions of myeloid DC as mean fluorescence intensity (MFI) were significantly less in maternal blood and in cord blood than in control (P<0.001, respectively). The DC differentiation factors, TNF-alpha and GM-CSF, released from mononuclear cells after lipopolysaccharide stimulation were significantly lower in maternal blood than in control (P<0.01). The distribution of DC subtypes was different in maternal and cord blood from those of non-pregnant women. Their role during pregnancy remains to be determined.

Cross-protective efficacy of the O1 Manisa + O 3039 bivalent vaccine and the O 3039 monovalent vaccine against heterologous challenge with FMDV O/Jincheon/SKR/2014 in pig.

After massive foot-and-mouth disease (FMD) outbreaks originated from Jincheon County from Dec. 2014 to Apr. 2015, the effectiveness of the previous FMD vaccine containing only the O1 Manisa as the O antigen, O1 Manisa + A Malaysia 97 + Asia 1 Sharmir trivalent vaccine, was questioned in South Korea, and a change in the O antigen in FMD vaccines was demanded to control the FMD caused by FMDV O/Jincheon/SKR/2014, the O Jincheon strain. Therefore, the efficacies of O1 Manisa + O 3039 bivalent vaccine and O 3039 monovalent vaccine were studied for cross-protection against heterologous challenge with the O Jincheon strain. In this study, the efficacy of the O1 Manisa + O 3039 bivalent vaccine was better than that of the O 3039 monovalent vaccine, even though the serological relationship (r1 value) between O Jincheon and O 3039 was matched according to the OIE Terrestrial Manual. According to serological test results from vaccinated specific pathogen free pigs, virus neutralization test titers against Jincheon were good estimates for predicting protection against challenge. A field trial of the O1 Manisa + O 3039 bivalent vaccine was performed to estimate the possibility of field application in conventional pig farms, especially due to concerns about the effect of maternally derived antibodies (MDA) in field application of the FMD vaccine. According to the result of the field trial, the O1 Manisa + O 3039 bivalent vaccine was considered to overcome MDA. The results of the efficacy and field trials indicated that the O1 Manisa + O3039 vaccine could be suitable to replace previous FMD vaccines to control the FMD field situation caused by O Jincheon FMDV.

Unique expression of a small IL-32 protein in the Jurkat leukemic T cell line.

Interleukin (IL)-32 was recently identified as a new cytokine which induces various proinflammatory cytokines in human monocytes and macrophages. Therefore, IL-32 has been primarily studied in inflammatory models such as rheumatoid arthritis and inflammatory bowel diseases. The regulation of endogenous IL-32 in other immune cells remains unknown. In the present study, we stimulated Jurkat T cells with phytohaemagglutinin (PHA) and phorbol myristate acetate (PMA) and examined IL-32 expression at both the mRNA and protein levels. All mRNAs of the four IL-32 isoforms and the 12-15 kDa IL-32 protein were independent of PHA and PMA stimulation, however a 9 kDa molecular weight IL-32 protein in the cell culture supernatant was induced by PHA and PMA after 16 h of stimulation. Compared to other human cell lines, the Jurkat cell line constitutively expressed a 12-15 kDa molecule of IL-32, which is smaller than the known IL-32 isoforms. We used IL-32 shRNA to examine the specificity of the 12-15 kDa molecule. Upon IL-32 shRNA transfection, the 12-15 kDa band was decreased specifically as compared to the control scrambled clone. Thus, the constitutive expression of IL-32 mRNA as well as the predominant production of a smaller sized IL-32 isoform in Jurkat cells may implicate a role for IL-32 in human T cell leukemia.