Technetium (99mTc) exametazime (hexamethylpropyleneamine oxime, HMPAO) labeled leukocyte scintigraphy is mainly used to exclude occult infection in our institution. On review of previously published article, no case of popliteal venous aneurysm was ever diagnosed and detected on labeled leukocyte scintigraphy. We present a rare case of popliteal venous aneurysm which was detected on labeled leukocyte scintigraphy and was further confirmed with single-photon emission computed tomography and computed tomography fusion.
BACKGROUND:
Feline Infectious Peritonitis (FIP) is a lethal systemic disease, caused by the FIP Virus (FIPV); a virulent mutant of Feline Enteric Coronavirus (FECV). Currently, the viruses virulence determinants and host gene expressions during FIPV infection are not fully understood.
METHODS:
RNA sequencing of Crandell Rees Feline Kidney (CRFK) cells, infected with FIPV strain 79-1146 at 3 hours post infection (h.p.i), were sequenced using the Illumina next generation sequencing approach. Bioinformatic's analysis, based on Felis catus 2X annotated shotgun reference genome, using CLC bio Genome Workbench mapped both control and infected cell reads to 18899 genes out of 19046 annotated genes. Kal's Z test statistical analysis was used to analyse the differentially expressed genes from the infected CRFK cells. Real time RT-qPCR was developed for further transcriptional profiling of three genes (PD-1, PD-L1 and A3H) in infected CRFK cells and Peripheral Blood Mononuclear Cells (PBMCs) from healthy and FIP-diseased cats.
RESULTS:
Based on Kal's Z-test, with False Discovery Rate (FDR) <0.05 and >1.99 fold change on gene expressions, a total of 61 genes were differentially expressed by both samples, where 44 genes were up-regulated and the remainder were down-regulated. Most genes were closely clustered together, suggesting a homogeneous expression. The majority of the genes that were significantly regulated, were those associated with monocytes-macrophage and Th1 cell functions, and the regulation of apoptosis. Real time RT-qPCR developed focusing on 2 up-regulated genes (PD-L1 and A3H) together with an apoptosis associated gene PD-1 expressions in FIPV infected CRFK cells and in PBMCs from healthy and FIP diagnosed cats produced concordant results with transcriptome data.
CONCLUSION:
The possible roles of these genes, and their importance in feline coronaviruses infection, are discussed.
Hepatitis B virus (HBV) is a noncytopathic virus and billions of HBV-infected patients live uneventful lives and do not suffer from notable liver damage. However, HBV also causes progressive liver diseases characterized by hepatic inflammation, hepatic fibrosis, and liver cancer in millions of HBV-infected patients. The goal of this study was to evaluate the role of mutant HBV in HBV pathogenesis. In a cohort of 360 chronic HBV-infected patients, mutations at T1762/A1764 of HBV genome were detected in most of the patients with HBV-induced liver cirrhosis and hepatocellular carcinoma. To explore if mutations at T1762/A1764 of HBV genome has any role in progressive liver disease, peripheral blood mononuclear cells (PBMCs) and antigen-presenting dendritic cells (DCs) were isolated from five chronic hepatitis B (CHB) patients with mutations at T1762/A1764 and five comparable patients of CHB without mutations at T1762/A1764. DCs were pulsed with hepatitis B surface antigen (HBsAg). The levels of cytokines produced by PBMCs and DCs as well as nitrite production by DCs were evaluated. Significantly higher levels of interleukin-12, tumor necrosis factor-alpha, interferon-gamma, and transforming growth factor-beta were detected in cultures of PBMCs, DCs, and HBsAg-pulsed DCs from CHB patients with mutations at T1762/A1764 compared with those without mutations (p
Research on natural products has been widely used as a strategy to discover new drugs with potential for applications in complementary medicines because they have fewer side effects than conventional drugs. The aim of the present study was to evaluate the in vitro cytotoxic effects of crude aqueous Catharanthus roseus extract on Jurkat cells and normal peripheral blood mononuclear cells (PBMCs). The aqueous extract was standardised to vinblastine by high performance liquid chromatography (HPLC) and was used to determine cytotoxicity by the MTS [3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. DNA fragmentation assay was employed to determine if cell death was due to apoptosis. The results showed that the aqueous extract induced cell death of Jurkat cells at 24, 48 and 72 hours post-treatment in a time- and dose-dependent manner. However, cells treated at 48 and 72 hours produced higher cytotoxic effects with half maximal inhibitory concentration (IC50) values of 2.55 μg/ml and 2.38 μg/ml, respectively. In contrast, the extract induced normal PBMC proliferation, especially after 24 hours treatment with 1000 μg/ml. This result indicates that the C. roseus crude aqueous extract showed differential effects of inhibiting the proliferation of the Jurkat cell line and promoting the growth of PBMCs. These data suggest that the extract may be applicable for modulating the normal and transformed immune cells in leukaemia patients.
Research on natural products has been widely used as a strategy to discover new drugs with potential for applications in complementary medicines because they have fewer side effects than conventional drugs. The aim of the present study was to evaluate the in vitro cytotoxic effects of crude aqueous Catharanthus roseus extract on Jurkat cells and normal peripheral blood mononuclear cells (PBMCs). The aqueous extract was
standardised to vinblastine by high performance liquid chromatography (HPLC) and was used to determine cytotoxicity by the MTS [3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. DNA fragmentation assay was employed to determine if cell death was due to apoptosis. The results showed that the aqueous extract induced cell death of Jurkat cells at 24, 48 and 72 hours posttreatment in a time- and dose-dependent manner. However, cells treated at 48 and 72 hours produced higher cytotoxic effects with half maximal inhibitory concentration (IC50)values of 2.55 µg/ml and 2.38 µg/ml, respectively. In contrast, the extract induced normal PBMC proliferation, especially after 24 hours treatment with 1000 µg/ml. This result indicates that the C. roseus crude aqueous extract showed differential effects of inhibiting the proliferation of the Jurkat cell line and promoting the growth of PBMCs. These data suggest that the extract may be applicable for modulating the normal and transformed immune cells in leukaemia patients.
Increased susceptibility of diabetics to melioidosis, a disease caused by the Burkholderia pseudomallei bacterium is believed to be attributed to dysfunction of the innate immune system. However, the underlying mechanism of the innate susceptibility is not well-understood. Glycogen synthase kinase-3β (GSK3β) plays an important role in the innate inflammatory response caused by bacterial pathogens. The present study was conducted to investigate the effects of GSK3β inhibition by LiCl on levels of pro- and anti-inflammatory cytokines; and the activity of transcription factor NF-κB in B. pseudomallei-infected peripheral blood mononuclear cells (PBMC) derived from diabetic-induced and normal Sprague Dawley rats. In addition, the effects of LiCl on intracellular bacterial counts were also investigated. Infection of PBMC from diabetic and normal rats with B. pseudomallei resulted in elevated levels of cytokines (TNF-α, IL-12 and IL-10) and phosphorylation of NF-κB in both cell types. Intracellular bacterial counts decreased with time in both cell types during infection. However bacterial clearance was less prominent in diabetic PBMC. Burkholderia pseudomallei infection also caused inactivation (Ser9 phosphorylation) of GSK3β in normal PBMC, an effect absent in infected diabetic PBMC. Inhibition of GSK3β by LiCl lowered the levels of pro-inflammatory cytokines (TNF-α and IL-12) in both normal and diabetic PBMC. Similarly, phosphorylated NF- κB (pNF-κB) levels in both cell types were decreased with LiCl treatment. Also, LiCl was able to significantly decrease the intracellular bacterial count in normal as well as diabetic PBMC. Interestingly, the levels of anti-inflammatory cytokine IL-10 in both normal and diabetic PBMC were further elevated with GSK3β inhibition. More importantly, GSK3β in infected diabetic PBMC was inactivated as in their non-diabetic counterparts upon LiCl treatment. Taken together, our results suggest that inhibition of dysregulated GSK3β in diabetic PBMC resulted in the inactivation of NF-κB and modulation of inflammatory cytokine levels. This is evidence that dysregulation of GSK3β is a contributing factor in the molecular basis of innate dysfunction and susceptibility of diabetic host to melioidosis infection.
The purification of parasite-infected erythrocytes from whole blood containing leucocytes is crucial for many downstream genetic and molecular assays in parasitology. Current methodologies to achieve this are often costly and time consuming. Here, we demonstrate the successful application of a cheap and simple Non-Woven Fabric (NWF) filter for the purification of parasitized red blood cells from whole blood. NWF filtration was applied to the malaria-parasitized blood of three strains of mice, and one strain of rat, and to Babesia gibsoni parasitized dog blood. Before and after filtration, the white blood cell (WBC) removal rates and red blood cell (RBC) recovery rates were measured. After NWF filter treatment of rodent malaria-infected blood, the WBC removal rates and RBC recovery rates were, for Kunming mice: 99.51%±0.30% and 86.12%±8.37%; for BALB/C mice: 99.61%±0.15% and 80.74%±7.11%; for C57 mice: 99.71%±0.12% and 84.87%±3.83%; for Sprague-Dawley rats: 99.93%±0.03% and 83.30%±2.96%. Microscopy showed WBCs were efficiently removed from infected dog blood samples, and there was no obvious morphological change of B. gibsoni parasites. NWF filters efficiently remove leukocytes from malaria parasite-infected mouse and rat blood, and are also suitable for filtration of B. gibsoni-infected dog blood.
Acute myeloid leukemia (AML) is a malignant disease progressed from abnormal production of immature myeloid cells, which is often associated with concurrent infections after diagnosis. It was widely established that infections are the major contributors to mortality in this group due to the prevalency of neutropenia. Gram-negative Burkholderia pseudomallei is the causative agent of melioidosis. This disease had been reported in several neutropenic cancer patients undergoing chemotherapy resulting in severe clinical presentations and high mortalities which is in need of critical attention. Studies show that cytokines are important mediators of melioidosis progression and low neutrophil counts are associated with progression of its severity. However, to date, there are no reports on cytokine production in neutropenic cancer patients who are prone to melioidosis. Hence, here we assessed the cytokine production in neutropenic AML patients by introducing B. pseudomallei to their peripheral blood mononuclear cell (PBMC) culture in vitro. We observed that inflammatory response related cytokines namely TNF-α, IFN-γ IL-6 and IL-10 were highly circulated in infected PBMCs suggesting that these cytokines may play important roles in the progression of severity in melioidosis infected neutropenic patients.
Crimean-Congo haemorrhagic fever (CCHF) is a severe human infection which can lead to fatal consequences. Acute CCHF patients were previously shown to exhibit frequencies of regulatory T-cell (Treg) but lower Treg-mediated suppressive activities than the healthy counterparts. This study aims is to investigate the phosphorylation levels of Foxp3 protein (master regulator of Treg cells) in CCHF patients. Blood samples collected from 18 CCHF patients and nine healthy volunteers were used to isolate peripheral blood mononuclear cells (PBMCs). Total and phosphorylated Foxp3 expression levels in the isolated PBMC samples were monitored by western blot and quantified using ImageJ software. Total Foxp3 expression levels in CCHF patients displayed decreasing trend, but not significantly. In contrast, significantly lower expression levels of phosphorylated Foxp3 were reported in CCHF patients. Our results suggest a possible association between Foxp3 dephosphorylation and CCHF pathogenesis. Nevertheless, more studies are required to evaluate the effect of Foxp3 dephosphorylation on Treg function, which would not only help to enlighten the CCHF pathogenesis but also contribute to the development of effective treatment strategies.
The caspase inhibitor benzyloxycarbony (Cbz)-l-Val-Ala-Asp (OMe)-fluoromethylketone (z-VAD-FMK) has recently been shown to inhibit T cell proliferation without blocking caspase-8 and caspase-3 activation in primary T cells. We showed in this study that z-VAD-FMK treatment leads to a decrease in intracellular glutathione (GSH) with a concomitant increase in reactive oxygen species (ROS) levels in activated T cells. The inhibition of anti-CD3-mediated T cell proliferation induced by z-VAD-FMK was abolished by the presence of low molecular weight thiols such as GSH, N-acetylcysteine (NAC) and l-cysteine, whereas d-cysteine which cannot be metabolised to GSH has no effect. These results suggest that the depletion of intracellular GSH is the underlying cause of z-VAD-FMK-mediated inhibition of T cell activation and proliferation. The presence of exogenous GSH also attenuated the inhibition of anti-CD3-induced CD25 and CD69 expression mediated by z-VAD-FMK. However, none of the low molecular weight thiols were able to restore the caspase-inhibitory properties of z-VAD-FMK in activated T cells where caspase-8 and caspase-3 remain activated and processed into their respective subunits in the presence of the caspase inhibitor. This suggests that the inhibition of T cell proliferation can be uncoupled from the caspase-inhibitory properties of z-VAD-FMK. Taken together, the immunosuppressive effects in primary T cells mediated by z-VAD-FMK are due to oxidative stress via the depletion of GSH.
This is the first report of high-resolution human leukocyte antigen (HLA) typing in four indigenous groups in Malaysia. A total of 99 normal, healthy participants representing the Negrito (Jehai and Kensiu), Proto-Malay (Temuan) and a native group of Borneo (Bidayuh) were typed for HLA-A, -B, -DRB1 and -DQB1 genes using sequence-based typing. Eleven HLA-A, 26 HLA-B, 16 HLA-DRB1 and 14 HLA-DQB1 alleles were detected, including a new allele, HLA-B*3589 in the Jehai. Highly frequent alleles were A*2407, B*1513, B*1801, DRB1*0901, DRB1*1202, DRB1*1502, DQB1*0303 and DQB1*0502. Principal component analysis based on high-resolution HLA-A, -B and -DRB1 allele frequencies showed close affinities among all four groups, including the Negritos, with other Southeast Asian populations. These results showed the scope of HLA diversity in these indigenous minority groups and may prove beneficial for future disease association, anthropological and forensic studies.
Leukaemia inhibitory factor (LIF) plays an indispensible role in embryo implantation. Aberrant LIF production is linked to implantation failure. LIF regulates multiple processes prior to and during implantation such as uterine transformation into a receptive state, decidualization, blastocyst growth and development, embryo-endometrial interaction, trophoblast invasion, and immune modulation. Due to its critical role, LIF has been a target for a nonhormonal contraception. In this review, we summarize up-to-date information on the role of LIF in implantation and its role in contraception.
Prostaglandins (PGs), derivatives of arachidonic acid, play an indispensable role in embryo implantation. PGs have been reported to participate in the increase in vascular permeability, stromal decidualization, blastocyst growth and development, leukocyte recruitment, embryo transport, trophoblast invasion, and extracellular matrix remodeling during implantation. Deranged PGs syntheses and actions will result in implantation failure. This review summarizes up-to-date literatures on the role of PGs in blastocyst implantation which could provide a broad perspective to guide further research in this field.
We determined the differential expression levels of proteins in peripheral blood mononuclear cells of patients with dengue fever (DF) and dengue hemorrhagic fever (DHF). Proteins were subjected to two-dimensional electrophoresis, mass spectrometry and Western blot analysis. We identified 8 proteins that were 2-fold or more up-regulated in patients compared to healthy control, three of which, aldolase, thioredoxin peroxidase and alpha tubulin, were related to dengue infection. Both thioredoxin peroxidase and alpha tubulin were over-expressed 4.9 and 3.3 times respectively in DHF compared to DF patients while aldolase was up-regulated 2.2 times in DF compared to DHF patients. Alpha tubulin and thioredoxin peroxidase have the potential to be utilized as biomarkers for DHF.
The occurrence of adult Gnathostoma malaysiae in Rattus surifer and R. tiomanicus in Malaysia has been reported but there are no known reports on the host tissue reactions. This paper reports on the gross pathology caused by G. malaysiae in a red spiny forest rat, R. surifer and the tissue reactions caused. A tumor-like growth was located on the mid-stomach wall in a female rat captured in Gunung Bachock, Kelantan, Malaysia. This growth consisted of four tunnel-like structures containing sanguinopurulent fluid and leukocytes and this structure led into a central canal. The tissue surrounding the tumor was greatly inflamed and there was localized gastritis. The tunnel-like structure was surrounded by dense fibrotic tissue. The stomach wall was devoid of superficial epithelium and smooth muscle but mucinous glands were present. The midregion of the fibrotic scar contained eggs of G. malaysiae which had evoked a strong tissue reaction and were surrounded by pus. Blood vessels were empty, dilated and had undergone vasculitis and thrombosis.
Aqueous extract of Ficus deltoidea var. agustifolia was examined for the subchronic toxicity effects in rats. Groups of 10 rats were given the extract daily by oral gavage for 90 days at 0 (control), 100 and 300mg/kg/body weight, respectively. Blood samples were collected upon sacrificed and analysed for haemogram and biochemistry. The results showed there were no significant changes of the blood parameters in all treated groups compared to the control.
Tuberculosis and malignancy are two common causes of exudative pleural effusions. In this retrospective study of 52 patients with tuberculous pleural effusions and 32 patients with malignant effusions, the median age of patients with malignant effusions (68.5 years) was older than that of patients with tuberculous effusions (34.5 years) (p < 0.001). Both types of effusion occurred more frequently on the right side and there was no difference between them in terms of right-sided dominance. A higher percentage of patients with malignant pleural effusions (44%) presented with large effusions than patients with tuberculous effusions (12%) (x2 = 11.33, p = 0.001). A higher proportion of patients with tuberculous effusion had lymphocyte predominant effusions and tuberculous effusions had higher lymphocyte percentage, lower red cell count, and higher protein content. However, there was considerable overlap of these characteristics of both types of effusions.