Role of immune activation in CD4<Superscript>+</Superscript> T-cell depletion in HIV-1 infected Indian patients

Objectives

The correlation of immune activation with CD4⁺ depletion and HIV-1 disease progression has been evidenced by several studies involving mainly clade B virus. However, this needs to be investigated in developing countries such as India predominately infected with clade C virus.

Materials and methods

In a cross-sectional study of 68 antiretroviral treatment naïve, HIV-1 infected Indian patients, we studied the association between CD4⁺ T cells, plasma HIV-1 RNA levels, and immune activation markers using unadjusted and adjusted correlative analyses.

Results

Significant negative correlations of higher magnitude were observed between the CD4⁺ T cell percentages and plasma HIV-1 RNA levels in the study population when adjusted for the effects of immune activation markers. However, the negative association of CD4⁺ T cells with immune activation markers remained unaffected when controlled for the effects of plasma HIV-1 RNA levels.

Conclusions

Our results support the important role of immune activation in CD4⁺ T cell depletion and disease progression during untreated HIV-1 infection.

     

A CD57<Superscript>+</Superscript> CTL Degranulation Assay Effectively Identifies Familial Hemophagocytic Lymphohistiocytosis Type 3 Patients

Purpose

Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) is a genetic disorder that results in immune dysregulation. It requires prompt and accurate diagnosis. A natural killer (NK) cell degranulation assay is often used to screen for FHL3 patients. However, we recently encountered two cases of late-onset FHL3 carrying novel UNC13D missense mutations: in these cases, the degranulation assays using freshly isolated and interleukin (IL)-2-activated NK cells yielded contradictory results. Since the defective degranulation of CD57⁺ cytotoxic T lymphocytes (CTLs) in these cases was helpful for making the diagnosis, we assessed whether the CD57⁺ CTL degranulation assay more effectively identified FHL3 patients than the NK cell assays.

Methods

Forty additional patients with hemophagocytic lymphohistiocytosis were prospectively screened for FHL3 by measuring the perforin expression in NK cells and the expression of Munc13-4, syntaxin-11, and Munc18-2 in platelets and by performing NK cell and CTL degranulation assays. The results were confirmed by genetic analysis.

Results

The freshly isolated NK cell degranulation assay detected FHL3 patients with high sensitivity (100%) but low specificity (71%). The IL-2-stimulated NK cell assay had improved specificity, but 3 out of the 31 non-FHL3 patients still showed degranulation below the threshold level. The CD57⁺ CTL degranulation assay identified FHL3 patients with high sensitivity and specificity (both 100%).

Conclusions

The CD57⁺ CTL degranulation assay more effectively identified FHL3 patients than the NK cell-based assays.

     

Greater frequency of CD5-negative CD8<Superscript>+</Superscript> T cells against human immunodeficiency virus type 1 than other viruses is consistent with adaptation to antigenic variation

Background

The CD5 protein antagonizes phosphorylation events downstream of T cell receptor (TCR) engagement to decrease T cell responsiveness. CD5-negative T cell clones respond preferentially over their CD5⁺ counterparts against cells with low human histocompatibility-linked leukocyte antigen (HLA) levels. In human immunodeficiency virus type 1 (HIV-1) infection, CD5^-CD8⁺ T cells increase in prevalence with disease progression.

Methods

To investigate potential causes of this expansion of CD5^-CD8⁺ T cells in HIV-1 infection, we compared CD5 expression on CD8⁺ T cells reactive against HIV-1 peptides, common viral peptides and a self peptide that together span a broad range of TCR avidities in the context of the common HLA-A2 class I restriction molecule. Following stimulation, CD5 expression on peptide-specific CD8⁺ T cells was assessed by flow cytometry.

Results

In healthy controls, there was no significant difference in the CD5⁺ percentage of CD8⁺ T cells specific for common viral peptides, but a lower percentage of those responding against a common self peptide expressed CD5. The same relationship occurred in HIV-infected individuals, however, a lower percentage of HIV peptide-specific CD8⁺ T cells than other viral peptide-specific CD8⁺ T cells expressed CD5. In terms of overall CD5 expression level at the peptide-specific responder population level, HIV-specific CD8⁺ T cells resembled those responsive against the self peptide, despite much higher avidity TCR/HLA/peptide interactions.

Conclusions

This deficit in CD5 expression selective for HIV-specific CD8⁺ T cells is consistent with in vivo adaptation to low avidity HIV peptide variants and has potential consequences for CD8⁺ T cell expansion, cross-reactivity and autoreactivity.

     

Varied sensitivity to therapy of HIV-1 strains in CD4<Superscript>+</Superscript> lymphocyte sub-populations upon ART initiation

Background

Although antiretroviral therapy (ART) has proven its success against HIV-1, the long lifespan of infected cells and viral latency prevent eradication. In this study we analyzed the sensitivity to ART of HIV-1 strains in naïve, central memory and effector memory CD4⁺ lymphocyte subsets.

Methods

From five patients cellular HIV-1 infection levels were quantified before and after initiation of therapy (2-5 weeks). Through sequencing the C2V3 region of the HIV-1 gp120 envelope, we studied the effect of short-term therapy on virus variants derived from naïve, central memory and effector memory CD4⁺ lymphocyte subsets.

Results

During short-term ART, HIV-1 infection levels declined in all lymphocyte subsets but not as much as RNA levels in serum. Virus diversity in the naïve and central memory lymphocyte populations remained unchanged, whilst diversity decreased in serum and the effector memory lymphocytes. ART differentially affected the virus populations co-circulating in one individual harboring a dual HIV-1 infection. Changes in V3 charge were found in all individuals after ART initiation with increases within the effector memory subset and decreases found in the naïve cell population.

Conclusions

During early ART virus diversity is affected mainly in the serum and effector memory cell compartments. Differential alterations in V3 charge were observed between effector memory and naïve populations. While certain cell populations can be targeted preferentially during early ART, some virus strains demonstrate varied sensitivity to therapy, as shown from studying two strains within a dual HIV-1 infected individual.

     

<Superscript>1</Superscript>H nuclear magnetic resonance (NMR)-based serum metabolomics of human gallbladder inflammation

Objective and design

We present in this article ¹H nuclear magnetic resonance (NMR)-based metabolic approach to screen the serum metabolic alterations in human gallbladder inflammation with chronic cholecystitis (CC).

Material/methods

Total of 71 human serum samples was divided into two groups, (n = 41, CC) and (n = 30 control). ¹H NMR metabolic profiling was carried out for investigation of metabolic alterations. Multivariate statistical analysis was applied for pattern recognition and identification of metabolites playing crucial role in gallbladder inflammation. Receiver operating curve (ROC) and pathway analysis on NMR data were also carried out to validate the findings.

Results

Serum metabolites such as glutamine, low-density lipoprotein (LDL), very low-density lipoprotein (VLDL), alanine, branch chained amino acids (BCAA), histidine and tyrosine were found to be depleted whereas formate, lactate, 1,2-propanediol were found to be elevated in CC. Metabolic pathways associated with metabolite alteration have also been reported.

Conclusions

NMR has been established for disease diagnosis along with identification of metabolic pattern recognition in biofluids. Gallstones cause inflammation of the gallbladder in the form of CC. Inflammation plays a major role in causation of gall bladder cancer and leads the way to malignancy. Metabolic analysis of CC may lead to early diagnosis of disease and its progression to gallbladder cancer.

     

Ouabain attenuates ovalbumin-induced airway inflammation

Purpose

Ouabain, an Na⁺/K⁺-ATPase inhibitor hormone, presents immunomodulatory actions, including anti-inflammatory effect on acute inflammation models.

Methods

In the present study, the effect of ouabain in a model of allergic airway inflammation induced by ovalbumin (OVA) was assessed.

Results

Initially, it was observed that ouabain treatment inhibited cellular migration induced by OVA on bronchoalveolar lavage fluid (BALF), mostly granulocytes, without modulating macrophage migration. In addition, it was observed, by flow cytometry, that ouabain reduces CD3^high lymphocytes cells on BALF. Furthermore, treatment with ouabain decreased IL-4 and IL-13 levels on BALF. Ouabain also promoted pulmonary histological alterations, including decreased cell migration into peribronchiolar and perivascular areas, and reduced mucus production in bronchioles regions observed through hematoxylin–eosin (HE) and by periodic acid-Schiff stain, respectively. Allergic airway inflammation is characterized by high OVA-specific IgE serum titer. This parameter was also reduced by the treatment with ouabain.

Conclusions

Therefore, our data demonstrate that ouabain negatively modulates allergic airway inflammation induced by OVA.

     

MiR-18a and miR-17 are positively correlated with circulating PD-1<Superscript>+</Superscript>ICOS<Superscript>+</Superscript> follicular helper T cells after hepatitis B vaccination in a chinese population

Background

While vaccination remains the most effective method to control hepatitis B virus (HBV) infection, 5–10% of recipients exhibit non-responsiveness to the HB vaccine. Immunological analysis of strong, weak or absent protective antibody responses to the HB vaccine should provide insights into the mechanisms that contribute to non-responsiveness.

Results

We investigated the potential involvement of follicular helper T (Tfh) cells in the immune response to HB vaccine, and associations between the miR-17–92 cluster and Tfh cells. We recruited 12 adults who had completed the HB vaccination course during childhood. Following a booster dose of HB vaccine, hepatitis B surface antibody (HBsAb) titers, percentage of PD-1+ICOS+ circulating Tfh (cTfh) and plasma cells, and expression of miR-17–92 were assessed at baseline (before immunization) and after vaccination on days 7 and 14. Notably, the HBsAb level gradually increased after HB vaccination while the proportion of PD-1+ICOS+ cTfh cells was significantly increased on day 7 relative to baseline, so as plasma cells. Expression of miR-18a and miR-17 within the miR-17–92 cluster and HBsAb titers in CD4+ T cells were positively correlated with the PD-1+ICOS+ cTfh cells proportions after HB vaccination.

Conclusions

The increase in HBsAb titers was positively associated with expression of all the components of the miR-17–92 cluster except miR-19a. Our findings indicate that the miR-17–92 cluster contributes to antibody production, and miR-18a and miR-17 are involved in Tfh cells differentiation after HB vaccination.

     

<Emphasis Type="Italic">Helicobacter bilis</Emphasis>-Associated Suppurative Cholangitis in a Patient with X-Linked Agammaglobulinemia

?

Helicobacter bilis is a commensal bacterium causing chronic hepatitis and colitis in mice. In humans, enterohepatic Helicobacter spp. are associated with chronic hepatobiliary diseases.

Purpose

We aimed at understanding the microbial etiology in a patient with X-linked agammaglobulinemia presenting with suppurative cholangitis.

Methods

16S rDNA PCR directly performed on a liver biopsy retrieved DNA of H. bilis.

Results

Clinical outcome resulted in the normalization of clinical and biological parameters under antibiotic treatment by a combination of ceftriaxone, metronidazole, and doxycyclin followed by a 2-week treatment with moxifloxacin and a 2-month treatment with azithromycin.

Conclusion

In conclusion, these data suggest a specific clinical and microbiological approach in patients with humoral deficiency in order to detect H. bilis hepatobiliary diseases.

     

The diagnostic performance of <Superscript>18</Superscript>F-FAMT PET and <Superscript>18</Superscript>F-FDG PET for malignancy detection: a meta-analysis

Background

This meta-analysis aims to compare the diagnostic performance of l-3-¹⁸F-α-methyl tyrosine (¹⁸F-FAMT) positron emission tomography (PET) and 2-deoxy-2-[¹⁸F]fluoro-d-glucose (¹⁸F-FDG) PET for malignancy detection.

Methods

The workflow of this study follows Cochrane Collaboration Guidelines of a systematic review of diagnostic test accuracy studies. An electronic search was performed for clinical diagnostic studies directly comparing ¹⁸F-FAMT and ¹⁸F-FDG PET for malignant tumors. Study quality, the risks of bias and sources of variation among studies were assessed using the QUADAS (Quality Assessment of Diagnostic Accuracy Studies) assessment tool. A separate meta-analysis was performed for diagnostic performance based on visual assessment and diagnostic cut-off values. Whenever possible, a bivariate random-effect model was used for analysis and pooling of diagnostic measures across studies.

Results

Electronic search revealed 56 peer-reviewed basic science investigations and clinical studies. Six eligible studies (272 patients) of various type of cancer were meta-analyzed. The ¹⁸F-FAMT diagnostic accuracy for malignancy was higher than ¹⁸F-FDG based on both visual assessment (diagnostic odd ratio (DOR): 8.90, 95% confidence interval (CI) [2.4, 32.5]) vs 4.63, 95% CI [1.8, 12.2], area under curve (AUC): 77.4% vs 72.8%) and diagnostic cut-off (DOR: 13.83, 95% CI [6.3, 30.6] vs 7.85, 95% CI [3.7, 16.8], AUC: 85.6% vs 80.2%), respectively. While the average sensitivity and specificity of ¹⁸F-FAMT and ¹⁸F-FDG based on visual assessment were similar, ¹⁸F-FAMT was significantly more specific than ¹⁸F-FDG (p?<?0.05) based on diagnostic cut-off values.

Conclusions

¹⁸F-FAMT is more specific for malignancy than ¹⁸F-FDG, while their sensitivity is comparable. ¹⁸F-FAMT PET is equal to ¹⁸F-FDG PET in diagnostic performance for malignancy detection in several cancer types.

     

Prevention of allergic airway hyperresponsiveness and remodeling in mice by <Emphasis Type="Italic">Astragaliradix</Emphasis> Antiasthmatic decoction

Background

Astragali radix Antiasthmatic Decoction (AAD), a traditional Chinese medication, is found effective in treating allergic diseases and chronic cough. The purpose of this study is to determine whether this medication could suppress allergen-induced airway hyperresponsiveness (AHR) and remodeling in mice, and its possible mechanisms.

Methods

A mouse model of chronic asthma was used to investigate the effects of AAD on the airway lesions. Mice were sensitized and challenged with ovalbumin (OVA), and the extent of AHR and airway remodeling were characterized. Cells and cytokines in the bronchoalveolar lavage fluid (BALF) were examined.

Results

AAD treatment effectively decreased OVA-induced AHR, eosinophilic airway inflammation, and collagen deposition around the airway. It significantly reduced the levels of IL-13 and TGF-β1, but exerted inconsiderable effect on INF-γ and IL-10.

Conclusions

AAD greatly improves the symptoms of allergic airway remodeling probably through inhibition of Th2 cytokines and TGF-β1.

     

<Emphasis Type="Italic">SERPING1</Emphasis> mRNA overexpression in monocytes from HIV+ patients

Objective

The HIV-1 virus activates the complement system, an essential element of the immune system. SERPING1 is a protease inhibitor that disables C1r/C1s in the C1 complex of the classical complement pathway.

Methods

In this paper, we performed an analysis of several microarrays deposited in GEO dataset to demonstrate that SERPING1 mRNA is modulated in CD14⁺ monocytes from HIV-1-infected individuals. In addition, data were validated on monocytes isolated from seronegative healthy volunteers, treated with IFNs.

Results

Our analysis shows that SERPING1 mRNA is overexpressed in monocytes from HIV-1+ patients and the expression levels correlate positively with viral load and negatively with the CD4⁺ T-cell count. Of note, anti-retroviral therapy is able to reduce the levels of SERPING1 mRNA, ex vivo. In addition, we found that 30% of the SERPING1 genes network is upregulated in monocytes from HIV-1+ patients. Noteworthy, the expression levels of IFITM1—an antiviral molecule belonging to the genes network—correlate positively with SERPING1 expression. Interestingly, the monocytes treatment with IFN-gamma, IFN-beta and IFN-alpha significantly upregulates the SERPING1 mRNA expression levels.

Conclusions

From the outcome of our investigation, it is possible to conclude that SERPING1 and its network serve as important components of the innate immune system to restrict HIV-1 infection.

     

Appearance of claudin-5<Superscript>+</Superscript> leukocytes in the central nervous system during neuroinflammation: a novel role for endothelial-derived extracellular vesicles

Background

The mechanism of leukocyte transendothelial migration (TEM) across the highly restrictive blood-brain barrier (BBB) remains enigmatic, with paracellular TEM thought to require leukocytes to somehow navigate the obstructive endothelial tight junctions (TJs). Transient interactions between TJ proteins on the respective leukocyte and endothelial surfaces have been proposed as one mechanism for TEM. Given the expanding role of extracellular vesicles (EVs) in intercellular communication, we investigated whether EVs derived from brain microvascular endothelial cells (BMEC) of the BBB may play a role in transferring a major TJ protein, claudin-5 (CLN-5), to leukocytes as a possible basis for such a mechanism during neuroinflammation.

Methods

High-resolution 3D confocal imaging was used to highlight CLN-5 immunoreactivity in the central nervous system (CNS) and on leukocytes of mice with the neuroinflammatory condition experimental autoimmune encephalomyelitis (EAE). Both Western blotting of circulating leukocytes from wild-type mice and fluorescence imaging of leukocyte-associated eGFP-CLN-5 in the blood and CNS of endothelial-targeted, Tie-2-eGFP-CLN-5 transgenic mice were used to confirm the presence of CLN-5 protein on these cells. EVs were isolated from TNF-α-stimulated BMEC cultures and blood plasma of Tie-2-eGFP-CLN-5 mice with EAE and evaluated for CLN-5 protein by Western blotting and fluorescence-activated cell sorting (FACS), respectively. Confocal imaging and FACS were used to detect binding of endothelial-derived EVs from these two sources to leukocytes in vitro. Serial electron microscopy (serial EM) and 3D contour-based surface reconstruction were employed to view EV-like structures at the leukocyte:BBB interface in situ in inflamed CNS microvessels.

Results

A subpopulation of leukocytes immunoreactive for CLN-5 on their surface was seen to infiltrate the CNS of mice with EAE and reside in close apposition to inflamed vessels. Confocal imaging of immunostained samples and Western blotting established the presence of CLN-5⁺ leukocytes in blood as well, implying these cells are present prior to TEM. Moreover, imaging of inflamed CNS vessels and the associated perivascular cell infiltrates from Tie-2-eGFP-CLN-5 mice with EAE revealed leukocytes bearing the eGFP label, further supporting the hypothesis CLN-5 is transferred from endothelial cells to circulating leukocytes in vivo. Western blotting of BMEC-derived EVs, corresponding in size to both exosomes and microvesicles, and FACS analysis of plasma-derived EVs from Tie-2-eGFP-CLN-5 mice with EAE validated expression of CLN-5 by EVs of endothelial origin. Confocal imaging and FACS further revealed both PKH-67-labeled EVs from cultured BMECs and eGFP-CLN-5⁺ EVs from plasma of Tie-2-eGFP-CLN-5 mice with EAE can bind to leukocytes. Lastly, serial EM and 3D contour-based surface reconstruction revealed a close association of EV-like structures between the marginating leukocytes and BMECs in situ during EAE.

Conclusions

During neuroinflammation, CLN-5⁺ leukocytes appear in the CNS, and both CLN-5⁺ leukocytes and CLN-5⁺ EVs are detected in the blood. As endothelial cells transfer CLN-5⁺ to leukocytes in vivo, and EVs released from BMEC bind to leukocytes in vitro, EVs may serve as the vehicles to transfer CLN-5 protein at sites of leukocyte:endothelial contact along the BBB. This action may be a prelude to facilitate TEM through the formation of temporary TJ protein bridges between these two cell types.

     

Novel Mutation in <Emphasis Type="Italic">CECR1</Emphasis> Leads to Deficiency of ADA2 with Associated Neutropenia

Purpose

Adenosine deaminase 2 (ADA2) have been reported to cause vasculitic diseases and immunodeficiency recently. Patients present with stroke episodes and rashes mimicking polyarteritis nodosa (PAN). We report a patient who has been followed up with severe neutropenia and found an unexpectedly revealed novel mutation in CECR1 affecting ADA2.

Methods

We reviewed medical records and clinical history of the patient. No mutations in other known neutropenia genes such as ELA, G6PC3, HAX1, AP3B1, LAMTOR2, VPS13B, VPS45, GFI1, JAGN1, or WAS could be detected. Sanger sequencing was used to confirm the genetic variants in the patient and relatives.

Results

Genetic analysis by exome sequencing revealed a novel mutation in the gene CECR1 (c.G962A; p.G321E) which segregated perfectly in the relatives.

Conclusion

This is the first DADA2 patient presenting with severe neutropenia. We suggest that in patients with unexplained cytopenias combined with immunodeficiency, fevers of unknown origin and high inflammation markers, DADA2 should be considered.

     

<Emphasis Type="Italic">Celastrus orbiculatus</Emphasis> extracts induce apoptosis in mTOR-overexpressed human hepatocellular carcinoma HepG2 cells

Background

Celastrus orbiculatus (Celastraceae) are used as traditional Chinese medicine to treat inflammation and cancer. This study aims to evaluate the effect of Celastrus orbiculatus extract (COE) on the apoptosis in human hepatic carcinoma HepG2 cells with mTOR overexpression.

Methods

The stable expression of mTOR in HepG2 cells (HepG2/mTOR⁺) were established by lipofectin transfection of GV238-mTOR recombinant plasmids and further antibiotic selection. Human hepatic carcinoma HepG2/mTOR⁺ cells were treated with different concentrations (20, 40, 80, 160, and 320?μg/mL) of COE for 24?h. The cell proliferation upon COE treatment was detected by MTT. Apoptosis was measured by Flow Cytometry. The activity of mTOR signaling pathway was detected by Western Blotting.

Results

COE significantly inhibited the proliferation of HepG2/mTOR⁺ cells. The expression levels of Bax and Caspase-3 protein were increased in the HepG2/mTOR⁺ cells in a dose-dependent manner. The proteins expression of Bcl2, Bcl-2?L12, mTOR, phospho-mTOR, 4EBP1, phospho-4EBP1, P70S6k, and phospho-P70S6k in HepG2/mTOR⁺ cells were reduced in dose-dependent manners. Furthermore, COE and mTOR inhibitor rapamycin (RAPA) synergistically induced apoptosis in HepG2/mTOR⁺ cells by regulating apoptosis-related proteins and inhibiting mTOR signaling pathways.

Conclusion

COE could inhibit the proliferation of HepG2/mTOR⁺ cells, and induce the cell apoptosis. The mechanisms may be related to the regulation of the expression of Bcl-2, Bcl-2?L12, and mTOR signaling pathways. These data suggest that COE may be a potential treatment for human hepatocellular carcinoma.

     

Heme oxygenase-1/biliverdin/carbon monoxide pathway downregulates hypernociception in rats by a mechanism dependent on cGMP/ATP-sensitive K<Superscript>+</Superscript> channels

Objective and design

To investigate the role of heme oxygenase-1 (HO-1), carbon monoxide (CO), and biliverdin (BVD) in the zymosan-induced TMJ arthritis in rats.

Materials and Methods

Mechanical threshold was assessed before and 4 h after TMJ arthritis induction in rats. Cell influx, myeloperoxidase activity, and histological changes were measured in the TMJ lavages and tissues. Trigeminal ganglion and periarticular tissues were used for HO-1, TNF-α, and IL-1β mRNA time course expression and immunohistochemical analyses. Hemin (0.1, 0.3, or 1 mg kg^?1), DMDC (0.025, 0.25, or 2.5 µmol kg^?1), biliverdin (1, 3, or 10 mg kg^?1), or ZnPP-IX (1, 3 or 9 mg kg^?1) were injected (s.c.) 60 min before zymosan. ODQ (12.5 µmol kg^?1; s.c.) or glibenclamide (10 mg kg^?1; i.p.) was administered 1 h and 30 min prior to DMDC (2.5 µmol kg^?1; s.c), respectively.

Results

Hemin (1 mg kg^?1), DMDC (2.5 µmol kg^?1), and BVD (10 mg kg^?1) reduced hypernociception and leukocyte migration, which ZnPP (3 mg kg^?1) enhanced. The effects of DMDC were counteracted by ODQ and glibenclamide. The HO-1, TNF-α, and IL-1β mRNA expression and immunolabelling increased.

Conclusions

HO-1/BVD/CO pathway activation provides anti-nociceptive and anti-inflammatory effects on the zymosan-induced TMJ hypernociception in rats.

     

Novel six-week protocol for generating functional human connective tissue-type (MC<Subscript>TC</Subscript>) mast cells from buffy coats

Objective

The aim of this study was to develop a novel protocol for generating large populations of fully mature and functional human mast cells (HMC) from CD34⁺ hematopoietic stem cells which require less culturing time than previously reported methods.

Methods

CD34⁺ cells isolated from fresh human buffy coats were sequentially cultured with different combinations of SCF, IL-6, IL-3, IL-9 and IL-4 under selected culturing conditions and time periods. Cells were then harvested for immunohistochemical characterization of morphological phenotypes and were functionally characterized by assessing their responses to IgE-dependent and -independent stimuli by measuring the release of inflammatory mediators and cytokines. Moreover, the pharmacological profiles of several classes of anti-inflammatory drugs in inhibiting the activation of these HMC were also characterized.

Results

We have developed a novel protocol that can generate large homogenous populations of mature and functional HMC in 6 weeks. These cells expressed both tryptase and chymase and were activated by anti-IgE, cationic peptides and calcium ionophores. Moreover, IgE-dependent activation of these cells was significantly inhibited by anti-inflammatory drugs. The morphological and functional characteristics of these mast cells resembled those of MC_TC type or connective tissue-type HMC.

Discussion

Our protocol represents a novel time-saving and economical approach for generating large numbers of primary HMC for functional studies of mast cell biology and for profiling novel anti-inflammatory therapeutic agents with mast cell-inhibitory properties in humans.

     

Inflammatory dysregulation of monocytes in pediatric patients with obsessive-compulsive disorder

Background

Although the exact etiology of obsessive-compulsive disorder (OCD) is unknown, there is growing evidence of a role for immune dysregulation in the pathophysiology of the disease, especially in the innate immune system including the microglia. To test this hypothesis, we studied inflammatory markers in monocytes from pediatric patients with OCD and from healthy controls.

Methods

We determined the percentages of total monocytes, CD16+ monocytes, and classical (CD14^highCD16?), intermediate (CD14^highCD16^low), and non-classical (CD14^lowCD16^high) monocyte subsets in 102 patients with early-onset OCD and in 47 healthy controls. Moreover, proinflammatory cytokine production (GM-CSF, IL-1β, IL-6, IL-8, and TNF-α) was measured by multiplex Luminex analysis in isolated monocyte cultures, in basal conditions, after exposure to lipopolysaccharide (LPS) to stimulate immune response or after exposure to LPS and the immunosuppressant dexamethasone.

Results

OCD patients had significantly higher percentages of total monocytes and CD16+ monocytes than healthy controls, mainly due to an increase in the intermediate subset but also in the non-classical monocytes. Monocytes from OCD patients released higher amounts of GM-CSF, IL-1β, IL-6, IL-8, and TNF-α than healthy controls after exposure to LPS. However, there were no significant differences in basal cytokine production or the sensitivity of monocytes to dexamethasone treatment between both groups. Based on monocyte subset distribution and cytokine production after LPS stimulation, patients receiving psychoactive medications seem to have an intermediate inflammatory profile, that is, lower than non-medicated OCD individuals and higher than healthy controls.

Conclusions

These results strongly support the involvement of an enhanced proinflammatory innate immune response in the etiopathogenesis of early-onset OCD.

     

Lesion based diagnostic performance of dual phase <Superscript>99m</Superscript>Tc-MIBI SPECT/CT imaging and ultrasonography in patients with secondary hyperparathyroidism

Background

We aimed to evaluate the diagnostic performance of ^99mTc-MIBI SPECT/CT and ultrasonography in patients with secondary hyperparathyroidism (SHPT), and explored the factors that affect the diagnostic performance.

Methods

^99mTc-MIBI SPECT/CT and ultrasonography were performed in 50 patients with SHPT within 1 month before they underwent surgery. Imaging results were confirmed by the pathology. Pearson correlation analysis was used to determine the correlation of PTH level with clinical data. The optimal cutoff value for predicting positive ^99mTc-MIBI results was evaluated by ROC analysis in lesions diameter.

Results

Forty-nine patients had a positive ^99mTc-MIBI imaging results and 39 patients had positive ultrasonography results. The sensitivities of ^99mTc-MIBI and ultrasonography were 98.00% and 78.00%, respectively. A total of 199 lesions were resected in 50 patients. Among them, 183 lesions were proved to be parathyroid hyperplasia. On per-lesion basis analysis, the sensitivity and specificity of ^99mTc-MIBI and ultrasonography were 59.34% and 75.00% vs 46.24% and 80.00%, respectively. The Pearson correlation analysis showed that the serum AKP and PTH level had a significant linear association (r?=?0.699, P?<?0.001). The lesion diameter was a statistically significant predictive factor in predicting positive ^99mTc-MIBI SPECT/CT. The optimal cutoff value for predicting positive ^99mTc-MIBI results evaluated by ROC analysis in lesions diameter was 8.05 mm.

Conclusion

Dual phase ^99mTc-MIBI SPECT/CT imaging had a higher sensitivity in patients with SHPT than ultrasonography. Therefore, using ^99mTc-MIBI positioning the lesion could be an effective method pre-surgical in patients with SHPT.