In Vitro Study of the Expression of CD1, CD14, CD25, CD30, CD35, CD95 Receptors by Macrophages of Mice Infected with Mycobacterium tuberculosis.

In cultures of peritoneal macrophages (MP) of male BALB/c mice infected with Mycobacterium tuberculosis from the BCG vaccine, the expression of CD1, CD14, CD25, CD30, CD35, and CD95 receptors was studied in vitro 3 months after infection. In MP cultures from intact and infected mice, mononuclear MP predominated (96 and 92%, respectively). Bi- and trinuclear MP in MP cultures from control and infected mice constituted 4 and 8.3% of all MP, respectively. In the cultures of both groups, no obvious correlations between the number of MP expressing CD-receptors and number of nuclei in these cells were found, but the expression of CD14 receptor was more often noted. In cultures from infected animals, hypertrophied MP and enhanced (by several times) expression of all CD-receptors were observed. The increase in the expression of CD-receptor can be determined by activation of plastic processes in hypertrophied MP (in epithelioid and in numerically insignificant polynuclear MP), which is due to the phenomenon of prolonged M. tuberculosis persistence in the vacuolar apparatus of these cells.

Effect of Acute and Prolonged Inflammation on the Gene Expression of Proinflammatory Cytokines and Their Receptors in the Anterior Pituitary Gland of Ewes.

An acute and prolonged inflammation inhibits the reproduction process by the disruption of the neurohormonal activity of the hypothalamic-pituitary-gonadal axis. It is thought that these changes may be caused by proinflammatory cytokines, i.e., interleukin (IL) -1ß, IL-6 and tumor necrosis factor (TNF) α. The aim of this study was to determine the effect of an acute and prolonged inflammation on the expression of genes encoding cytokine and their receptors, gonadotropin releasing hormone receptor (GnRHR), beta subunits of luteinizing hormone (LHß) and follicle-stimulating (FSHß) in the anterior pituitary (AP). Moreover, the circulating concentration of LH and FSH was also assayed. Two experiments were carried out on adult ewes which were divided into two control groups and treated with lipopolysaccharide (LPS; 400 ng / kg). Acute inflammation was caused by a single injection of LPS into the external jugular vein, while the chronic inflammation was induced by seven times LPS injection (one a day). In both experiments, animals were euthanized 3h after the last LPS / NaCl injection and the blood samples collected 15 min before euthanasia. An acute inflammation stimulates the expression of the IL-1ß, IL-6 and TNFα genes and their receptors in the AP of sheep. Prolonged inflammation increased TNFα gene expression and both types of TNFα and IL-6 receptors. Both an acute and prolonged inflammation inhibited LHß gene expression in the AP and reduced LH level in blood. A sevenfold LPS injection raises FSH concentration. The gene expression of GnRHR was reduced in the ovine AP only after a single injection of endotoxin. Our results suggest that there are important differences in the way how an acute and prolonged inflammation influence proinflammatory cytokines and their receptors gene expression in the AP of anestrous ewes, which could be reflected by differences in the AP secretory activity during these states.

Interleukin-35 Promotes Macrophage Survival and Improves Wound Healing After Myocardial Infarction in Mice.

RATIONALE: Targeting inflammation has been shown to provide clinical benefit in the field of cardiovascular diseases. Although manipulating regulatory T-cell function is an important goal of immunotherapy, the molecules that mediate their suppressive activity remain largely unknown. IL (interleukin)-35, an immunosuppressive cytokine mainly produced by regulatory T cells, is a novel member of the IL-12 family and is composed of an EBI3 (Epstein-Barr virus-induced gene 3) subunit and a p35 subunit. However, the role of IL-35 in infarct healing remains elusive. OBJECTIVE: This study aimed to determine whether IL-35 signaling is involved in healing and cardiac remodeling after myocardial infarction (MI) and, if so, to elucidate the underlying molecular mechanisms. METHODS AND RESULTS: IL-35 subunits (EBI3 and p35), which are mainly expressed in regulatory T cells, were upregulated in mice after MI. After IL-35 inhibition, mice showed impaired infarct healing and aggravated cardiac remodeling, as demonstrated by a significant increase in mortality because of cardiac rupture, decreased wall thickness, and worse cardiac function compared with wild-type MI mice. IL-35 inhibition also led to decreased expression of α-SMA (α-smooth muscle actin) and collagen I/III in the hearts of mice after MI. Pharmacological inhibition of IL-35 suppressed the accumulation of Ly6Clow and major histocompatibility complex IIlow/C-C motif chemokine receptor type 2- (MHC IIlow CCR2-) macrophages in infarcted hearts. IL-35 activated transcription of CX3CR1 (C-X3-C motif chemokine receptor 1) and TGF (transforming growth factor) ß1 in macrophages by inducing GP130 signaling, via IL12Rß2 and phosphorylation of STAT1 (signal transducer and activator of transcription family) and STAT4 and subsequently promoted Ly6Clow macrophage survival and extracellular matrix deposition. Moreover, compared with control MI mice, IL-35-treated MI mice showed increased expression of α-SMA and collagen within scars, correlating with decreased left ventricular rupture rates. CONCLUSIONS: IL-35 reduces cardiac rupture, improves wound healing, and attenuates cardiac remodeling after MI by promoting reparative CX3CR1+Ly6Clow macrophage survival.

Effects of inflammatory cytokines IFN-γ, TNF-α and IL-6 on the viability and functionality of human pluripotent stem cell-derived neural cells.

Multiple Sclerosis (MS) is an inflammatory neurodegenerative disease, where neural progenitor cell (NPC) transplantation has been suggested as a potential neuroprotective therapeutic strategy. Since the effect of inflammation on NPCs is poorly known, their effect on the survival and functionality of human NPCs were studied. Treatment with interleukin (IL)-6, tumor necrosis factor (TNF)-α and interferon (IFN)-γ did not induced cytotoxicity, but IFN-γ treatment showed decreased proliferation and neuronal migration. By contrast, increased proliferation and inhibition of electrical activity were detected after TNF-α treatment. Treatments induced secretion of inflammatory factors. Inflammatory cytokines appear to modulate proliferation as well as the cellular and functional properties of human NPCs.

Pituitary adenylate cyclase-activating polypeptide (PACAP-38) plays an inhibitory role against inflammation induced by chemical damage to zebrafish hair cells.

Pituitary adenylate cyclase-activating polypeptide (PACAP-38) is a common neuropeptide exerting a wide spectrum of functions in many fields, including immunology. In the present study, 5-day post-fertilization (dpf) zebrafish larvae of three diverse genetic lines [transgenic lines Tg(MPX:GFP) with GFP-labelled neutrophils and Tg(pou4f3:GAP-GFP) with GFP-labelled hair cells and the wild-type Tuebingen] were used to investigate an inhibitory role of PACAP-38 in inflammation associated with damaged hair cells of the lateral line. Individuals of each genetic line were assigned to four groups: (1) control, and those consisting of larvae exposed to (2) 10 µM CuSO4, (3) 10 µM CuSO4+100 nM PACAP-38 and (4) 100 nM PACAP-38, respectively. Forty-minute exposure to CuSO4 solution was applied to evoke necrosis of hair cells and consequent inflammation. The inhibitory role of PACAP-38 was investigated in vivo under a confocal microscope by counting neutrophils migrating towards damaged hair cells in Tg(MPX:GFP) larvae. In CuSO4-treated individuals, the number of neutrophils associated with hair cells was dramatically increased, while PACAP-38 co-treatment resulted in its over 2-fold decrease. However, co-treatment with PACAP-38 did not prevent hair cells from extensive necrosis, which was found in Tg(pou4f3:GAP-GFP) individuals. Real-Time PCR analysis performed in wild-type larvae demonstrated differential expression pattern of stress and inflammation inducible markers. The most significant findings showed that CuSO4 exposure up-regulated the expression of IL-8, IL-1ß, IL-6 and ATF3, while after PACAP-38 co-treatment expression levels of these genes were significantly decreased. The presence of transcripts for all PACAP receptors in neutrophils was also revealed. Adcyap1r1a and vipr1b appeared to be predominant forms. The present results suggest that PACAP-38 should be considered as a factor playing an important regulatory role in inflammatory response associated with pathological processes affecting zebrafish hair cells and it cannot be excluded that this interesting property has more universal significance.

Cutting Edge: Low-Affinity TCRs Support Regulatory T Cell Function in Autoimmunity.

Regulatory T cells (Tregs) use a distinct TCR repertoire and are more self-reactive compared with conventional T cells. However, the extent to which TCR affinity regulates the function of self-reactive Tregs is largely unknown. In this study, we used a two-TCR model to assess the role of TCR affinity in Treg function during autoimmunity. We observed that high- and low-affinity Tregs were recruited to the pancreas and contributed to protection from autoimmune diabetes. Interestingly, high-affinity cells preferentially upregulated the TCR-dependent Treg functional mediators IL-10, TIGIT, GITR, and CTLA4, whereas low-affinity cells displayed increased transcripts for Areg and Ebi3, suggesting distinct functional profiles. The results of this study suggest mechanistically distinct and potentially nonredundant roles for high- and low-affinity Tregs in controlling autoimmunity.

Mutations in TP53 and JAK2 are independent prognostic biomarkers in B-cell precursor acute lymphoblastic leukaemia.

BACKGROUND: In B-cell precursor acute lymphoblastic leukaemia (B-ALL), the identification of additional genetic alterations associated with poor prognosis is still of importance. We determined the frequency and prognostic impact of somatic mutations in children and adult cases with B-ALL treated with Spanish PETHEMA and SEHOP protocols. METHODS: Mutational status of hotspot regions of TP53, JAK2, PAX5, LEF1, CRLF2 and IL7R genes was determined by next-generation deep sequencing in 340 B-ALL patients (211 children and 129 adults). The associations between mutation status and clinicopathological features at the time of diagnosis, treatment outcome and survival were assessed. Univariate and multivariate survival analyses were performed to identify independent prognostic factors associated with overall survival (OS), event-free survival (EFS) and relapse rate (RR). RESULTS: A mutation rate of 12.4% was identified. The frequency of adult mutations was higher (20.2% vs 7.6%, P=0.001). TP53 was the most frequently mutated gene (4.1%), followed by JAK2 (3.8%), CRLF2 (2.9%), PAX5 (2.4%), LEF1 (0.6%) and IL7R (0.3%). All mutations were observed in B-ALL without ETV6-RUNX1 (P=0.047) or BCR-ABL1 fusions (P<0.0001). In children, TP53mut was associated with lower OS (5-year OS: 50% vs 86%, P=0.002) and EFS rates (5-year EFS: 50% vs 78.3%, P=0.009) and higher RR (5-year RR: 33.3% vs 18.6% P=0.037), and was independently associated with higher RR (hazard ratio (HR)=4.5; P=0.04). In adults, TP53mut was associated with a lower OS (5-year OS: 0% vs 43.3%, P=0.019) and a higher RR (5-year RR: 100% vs 61.4%, P=0.029), whereas JAK2mut was associated with a lower EFS (5-year EFS: 0% vs 30.6%, P=0.035) and a higher RR (5-year RR: 100% vs 60.4%, P=0.002). TP53mut was an independent risk factor for shorter OS (HR=2.3; P=0.035) and, together with JAK2mut, also were independent markers of poor prognosis for RR (TP53mut: HR=5.9; P=0.027 and JAK2mut: HR=5.6; P=0.036). CONCLUSIONS: TP53mut and JAK2mut are potential biomarkers associated with poor prognosis in B-ALL patients.

Molecular characterization of acute lymphoblastic leukemia with high CRLF2 gene expression in childhood.

BACKGROUND: A high-level expression of the CRLF2 gene is frequent in precursor B-cell acute lymphoblastic leukemia (pB-ALL) and can be caused by different genetic aberrations. The presence of the most frequent alteration, the P2RY8/CRLF2 fusion, was shown to be associated with a high relapse incidence in children treated according to ALL-Berlin-Frankfurt-Münster (BFM) protocols, which is poorly understood. Moreover, the frequency of other alterations has not been systematically analyzed yet. PROCEDURE: CRLF2 mRNA expression and potential genetic aberrations causing a CRLF2 high expression were prospectively assessed in 1,105 patients treated according to the Associazione Italiana Ematologia Oncologia Pediatrica (AIEOP)-BFM ALL 2009 protocol. Additionally, we determined copy number alterations in selected B-cell differentiation genes for all CRLF2 high-expressing pB-ALL cases, as well as JAK2 and CRLF2 mutations. RESULTS: A CRLF2 high expression was detected in 26/178 (15%) T-cell acute lymphoblastic leukemia (T-ALL) cases, 21 of them (81%) had been stratified as high-risk patients by treatment response. In pB-ALL, a CRLF2 high expression was determined in 91/927 (10%) cases; the P2RY8/CRLF2 rearrangement in 44/91 (48%) of them, supernumerary copies of CRLF2 in 18/91 (20%), and, notably, the IGH/CRLF2 translocation was detected in 16/91 (18%). Remarkably, 7 of 16 (44%) patients with IGH/CRLF2 translocation had already relapsed. P2RY8/CRLF2- and IGH/CRLF2-positive samples (70 and 94%, respectively) were characterized by a high frequency of additional deletions in B-cell differentiation genes such as IKZF1 or PAX5. CONCLUSION: Our data suggest that this high frequency of genetic aberrations in the context of a high CRLF2 expression could contribute to the high risk of relapse in P2RY8/CRLF2- and IGH/CRLF2-positive ALL.

Dendritic Cell-Mediated Mechanisms Triggered by LT-IIa-B5, a Mucosal Adjuvant Derived from a Type II Heat-Labile Enterotoxin of Escherichia coli.

Mucosal tissues are the initial site through which most pathogens invade. As such, vaccines and adjuvants that modulate mucosal immune functions have emerged as important agents for disease prevention. Herein, we investigated the immunomodulatory mechanisms of the B subunit of Escherichia coli heat-labile enterotoxin type IIa (LT-IIa-B5), a potent non-toxic mucosal adjuvant. Alternations in gene expression in response to LT-IIa-B5 were identified using a genome-wide transcriptional microarray that focused on dendritic cells (DC), a type of cell that broadly orchestrates adaptive and innate immune responses. We found that LT-IIa-B5 enhanced the homing capacity of DC into the lymph nodes and selectively regulated transcription of pro-inflammatory cytokines, chemokines, and cytokine receptors. These data are consistent with a model in which directional activation and differentiation of immune cells by LT-IIa-B5 serve as a critical mechanism whereby this potent adjuvant amplifies mucosal immunity to co-administered antigens.

CRLF2 overexpression identifies an unfavourable subgroup of adult B-cell precursor acute lymphoblastic leukemia lacking recurrent genetic abnormalities.

BACKGROUND: A deregulated CRLF2 (d-CRLF2) expression was described in B-cell acute lymphoblastic leukemia without recurrent fusion genes (B-NEG ALL). While the role of d-CRLF2 in children has been extensively described, little is known about its role and impact in adult ALL. METHODS: Expression levels of CRLF2 were evaluated by quantitative real-time PCR in 102 newly-diagnosed adult B-NEG ALL and correlated with the clinico-biological characteristics and outcome. Incidence and clinical impact of the P2RY8/CRLF2 transcript was also assessed. RESULTS: High CRLF2 levels, as continuous variable, were significantly associated with hyperleucocytosis (p=0.0002) and thrombocytopenia (p=0.005); when a cut-point at ΔCt≤8 was applied, 35 cases (34.3%), mostly males (80%), proved positive for CRLF2 expression. High CRLF2 levels, as continuous or categorical variable, were associated with a worse disease-free (p=0.003 and p=0.015) and overall survival (p=0.017 and 0.0038). Furthermore, when CRLF2 was analyzed as a categorical variable, a high statistical association was found with IKZF1 deletion and mutations in the JAK/STAT pathway (p=0.001 and p<0.0001, respectively). Finally, the P2RY8/CRLF2 transcript, identified in 8/102 patients (7.8%), was associated with a poor outcome. CONCLUSIONS: In adult B-NEG ALL, high CRLF2 expression is associated with distinct clinico-biological features and an unfavourable prognosis in both univariate and multivariate analysis; similarly, P2RY8/CRLF2 positivity correlates with a poor outcome. The quantification of CRLF2 is an important prognostic marker in adult B-lineage ALL without known genetic lesions.

A Potential Mechanism of High-Dose Ticagrelor in Modulating Platelet Activity and Atherosclerosis Mediated by Thymic Stromal Lymphopoietin Receptor.

Abnormal expression of thymic stromal lymphopoietin (TSLP) and its receptor (TSLPR) was found in patients with acute coronary syndrome. Ticagrelor, an oral platelet ADP P2Y12 receptor antagonist, is widely used in these patients. The aim of this study was to verify whether different doses of ticagrelor regulated plaque progression and platelet activity by modulating TSLP/TSLPR. Seventy-five ApoE-/- mice were randomly divided into five groups: (1) high-cholesterol diet (HCD, n = 15); (2) HCD plus ticagrelor 25 mg/kg/d (T1, n = 15); (3) HCD plus ticagrelor 50 mg/kg/d (T2, n = 15); (4) HCD plus ticagrelor 100 mg/kg/d (T3, n = 15); and (5) a normal diet group (ND, n = 15). At day 0 and at week 16, blood lipids and serum TSLP levels, expression of TSLPR, CD62, and CD63, platelet aggregation, platelet ATP release, PI3K/Akt signaling pathway, and plaque morphology were assessed. HCD increased TSLPR expression and atherosclerosis progression but high-dose ticagrelor (100 mg/kg) moderated this trend. TSLPR was positively correlated with Akt1, platelet aggregation, corrected plaque area, and vulnerability index in the T3 group (P<0.01). In conclusion, low-dose ticagrelor only inhibited platelet activity. Besides this inhibition, high-dose ticagrelor modulated platelet activity and atherosclerosis mediated by TSLPR, potentially through the PI3K/Akt signal pathway.

Interleukin-27 induces the endothelial differentiation in Sca-1+ cardiac resident stem cells.

Cytokines play important roles in cardiac repair and regeneration. Recently, we demonstrated that interleukin (IL)-6 family cytokines induce the endothelial differentiation of Sca-1+ cardiac resident stem cells through STAT3/Pim-1 signaling pathway. In contrast, the biological functions of IL-12 family cytokines in heart remain to be elucidated, though they show structural homology with IL-6. In the present study, we examined the effects of IL-12 family cytokines on the transdifferentiation of cardiac Sca-1+ cells into cardiac cells. RT-PCR analyses revealed that IL-27 receptor α (IL-27Rα), but not IL-12R or IL-23R, was expressed in cardiac Sca-1+ cells. The transcript expression of IL-27 was elevated in murine hearts in cardiac injury models. Intriguingly, IL-27 stimulation for 14 days induced the endothelial cell (EC) marker genes, such as CD-31 and VE-cadherin. Immunoblot analyses clarified that IL-27 treatment rapidly phosphorylated STAT3. IL-27 upregulated the expression of Pim-1, but the overexpression of dominant negative STAT3 abrogated the induction of Pim-1 by IL-27. Finally, adenoviral transfection of dominant negative Pim-1 inhibited IL-27-induced EC differentiation of cardiac Sca-1+ cells. These findings demonstrated that IL-27 promoted the commitment of cardiac stem cells into the EC lineage, possibly leading to neovascularization as a novel biological function. IL-27 could not only regulate the inflammation but also contribute to the maintenance of the tissue homeostasis through stem cell differentiation at inflammatory sites.

Expression of Interferon Lambda 4 Is Associated with Reduced Proliferation and Increased Cell Death in Human Hepatic Cells.

Interferon lambda 4 (IFN-λ4) is a novel type-III interferon that can be generated only in individuals carrying a ΔG frame-shift allele of an exonic genetic variant (rs368234815-ΔG/TT). The rs368234815-ΔG allele is strongly associated with decreased clearance of hepatitis C virus (HCV) infection. Here, we further explored the biological function of IFN-λ4 expressed in human hepatic cells-a hepatoma cell line HepG2 and fresh primary human hepatocytes (PHHs). We performed live confocal imaging, cell death and proliferation assays, mRNA expression profiling, protein detection, and antibody blocking assays using transient and inducible stable in vitro systems. Not only did we observe significant intracellular retention of IFN-λ4 but also detected secreted IFN-λ4 in the culture media of expressing cells. Secreted IFN-λ4 induced strong activation of the interferon-stimulated genes (ISGs) in IFN-λ4-expressing and surrounding cells in transwell assays. Specifically, in PHHs, secreted IFN-λ4 induced expression of the CXCL10 transcript and a corresponding pro-inflammatory chemokine, IP-10. In IFN-λ4-expressing HepG2 cells, we also observed decreased proliferation and increased cell death. All IFN-λ4-induced phenotypes--activation of ISGs, decreased proliferation, and increased cell death--could be inhibited by an anti-IFN-λ4-specific antibody. Our study offers new insights into biology of IFN-λ4 and its possible role in HCV clearance.

Changes in Functional Activity of JEG-3 Trophoblast Cell Line in the Presence of Factors Secreted by Placenta.

BACKGROUND AND AIMS: Cells in the maternal-fetal interface secrete cytokines that regulate proliferation, migration, and trophoblast invasion during the first trimester of pregnancy and the limitation of these processes during the third trimester. The aim of the study was to evaluate the influence of factors secreted by human placenta during the first and third trimester of pregnancy on cytokine receptor expression and proliferative and migratory activity of JEG-3 trophoblast cells. METHODS: The research was conducted using the explant conditioned media of placentas obtained from healthy women with elective termination of pregnancy at 9-11 weeks and placentas of women whose pregnancy progressed without complications at 38-39 weeks. Assessment of surface molecule expression was performed using FACS Canto II flow cytometer (BD, USA). The proliferative activity of JEG-3 trophoblast cells was evaluated by dyeing with crystal violet vital dye. The migration activity of JEG-3 was evaluated using 24-well insert plates with polycarbonate inserts (pore size 8 microns). RESULTS: Expression of CD116, CD118, CD119, IFNγ-R2, CD120b, CD183, CD192, CD295, EGFR, and TGFß-R2 on JEG-3 was higher when the cells were incubated in the presence of the third trimester placental factors in comparison with the first trimester placental factors. Factors secreted by the placenta during the third trimester of pregnancy had more pronounced stimulatory effect on the proliferation and migration of trophoblast in comparison with baseline levels and with the effect of the first trimester placental factors. CONCLUSIONS: The findings suggest that the behavior of trophoblasts in vitro might not be representative of in vivo behavior in the absence of additional local factors that influence the trophoblast in vivo.

[Low expression of Tlr-1, -2, -4 a il-2, -2r, -10, -10r and TgFb1 in pseudosynovial tissue collected at revision total knee arthroplasty for aseptic loosening]. / Nízká exprese Tlr-1, -2, -4 a il-2, -2r, -10, -10r, TgFb1 v pseudosynoviální tkáni odebrané pri reoperaci asepticky uvolnené totální endoprotézy kycle a kolena.

PURPOSE OF THE STUDY: Aseptic loosening (AL) and periprosthetic osteolysis (PPOL) in total hip (THA) and knee (TKA) arthroplasty are linked to an inflammatory process initiated by wear debris released from artificial joints. There is still limited information about the contribution of Toll-like receptors (TLRs) and distinct regulatory cytokines to AL/PPOL in both joints. METHODS: In this study, we investigated mRNA expression of TLR-1,-2,-4 and cytokines/receptors (IL-2,-2R,-10,-10R, TGFb1) in pseudosynovial tissue obtained from 55 patients with aseptically failed THAs/TKAs and 37 control patients with hip/knee primary osteoarthritis (OA) using quantitative RT-PCR. Immunohistochemical staining was used to detect the corresponding proteins. Non-parametric Kruskal-Wallis and Mann-Whitney tests were used to determine differences between the patient groups. RESULTS: When comparing expression profiles between patients with aseptically failed THA and TKA, higher amounts of TLR-1,- 2,-4 and IL-2R mRNA transcripts were detected in THA patients. The mRNA expression of studied molecules (TLR-1,-2,-4, IL-2, IL-10, IL-2R, IL-10R, TGFb1) did not differ between THA and OA hip tissues. Lower mRNA expression of TLR-1,-2,- 4, IL-10, and IL-10R was detected in TKA when compared to control knee OA. Similar mRNA profiles of IL-2, IL-2R, and TGFb1 were observed in TKA and knee OA. Using immunohistochemistry, we detected low expression of TLR-1 protein in failed THA/TKA, whereas TLR-2 protein levels were higher in TKA/THA patients than in OA controls. High individual variability in TLR-4 protein levels was detected among patients with aseptically loosened THA and TKA. IL-10 protein levels were similar in THA and TKA patient subgroups and control subjects, whereas IL-10R protein level was higher in failed TKAs and OA controls than in THAs. No difference in IL-2 protein levels was detected between patients with THA/TKA and those with OA. DISCUSSION: Our data indicate close similarity between the expression patterns in aseptically failed THA and TKA. However, certain differences were observed which also suggest unique pathways associated with the end-stage of aseptic loosening in THA and TKA. For instance, differences in the size, shape and load of polyethylene particles between THA and TKA could play some role. The composition of THA and TKA and differences in terms of mechanical forces might also be involved. CONCLUSIONS: This is the fist study comparing the gene expression profile of a particular set of innate immunity regulatory molecules between tissues from aseptically failed THA and TKA. Low expression of TLR-1,-2,-4 and cytokines/receptors (IL-2, IL-2R, IL-10, IL-10R, and TGFb1) was observed in pseudosynovial tissues obtained from aseptically failed THAs and TKAs. Higher amount of TLR transcripts was detected in THA as compared to TKA. These findings indicate certain differences in the mechanism of aseptic loosening occurring at the site of THA and TKA. Further research is warranted.

Glucocorticoids regulate natural killer cell function epigenetically.

Although glucocorticoids are well known for their capacity to suppress the immune response, glucocorticoids can also promote immune responsiveness. It was the purpose of this investigation to evaluate the molecular basis for this apparent dichotomous immunologic effect. Glucocorticoid treatment of natural killer cells (NK) was shown to reduce NK cell cytolytic activity by reduction of histone promoter acetylation for perforin and granzyme B, which corresponded with reduced mRNA and protein for each. In contrast, glucocorticoid treatment increased histone acetylation at regulatory regions for interferon gamma and IL-6, as well as chromatin accessibility for each. This increase in histone acetylation was associated with increased proinflammatory cytokine mRNA and protein production upon cellular stimulation. These immunologic effects were evident at the level of the individual cell and demonstrate glucocorticoids to epigenetically reduce NK cell cytolytic activity while at the same time to prime NK cells for proinflammatory cytokine production.

Interferon lambda 1 expression in cervical cells differs between low-risk and high-risk human papillomavirus-positive women.

Persistent infection by high-risk (HR) human papillomavirus (HPV) types is a prerequisite for progression to cancer. HR-HPVs may lead to a deregulation of innate immunity by interfering with the epithelial type I interferon (IFN) response, whereas very little is known about type III IFNs, a key component of the mucosal antiviral response. This study reports a first attempt to evaluate the activation of type III IFN genes (IFN lambda 1-3), IFN lambda receptor genes (IFN-lambdaR1 and IL10R2), and IFN-induced genes (MxA, ISG15, ISG56) in HPV-positive and HPV-negative cervical cells from 154 women attending the gynecological unit of a university hospital in Rome. Despite an increased individual variability, a coordinated expression of several IFN lambda-related genes was observed. Furthermore, IFN lambda 1 and IFN-lambdaR1 genes were expressed at higher levels in cervical cells positive to low-risk (LR) HPV compared to HR-HPV and HPV-negative cells. Consistently, ISG15 expression was significantly higher in LR-HPV-infected women than in the other groups. Moreover, IFN lambda 1 expression decreased significantly with abnormal cytological results. This study is the first to show the activation of a type III IFN response in LR-HPV-positive cervical cells and suggests that the lack of this response in HR-HPV infection may be related to lesion progression.

Early-life growth hormone treatment to offspring of undernourished mothers alters metabolic parameters in primary adipocytes in adulthood.

Maternal undernutrition (UN) is associated with the development of obesity and metabolic complications in adult offspring. This study investigated the impact of preweaning growth hormone (GH) treatment on adipocyte functionality in adult male offspring. Sprague-Dawley rats were assigned either standard (C) or undernourished (UN) diet (50% ad libitum) throughout gestation. Postnatal day 3-21, male C/UN pups received either saline (CS, UNS) or GH (2.5 µg/g/d; CGH, UNGH) by subcutaneous injection. Primary adipocytes were isolated following the collagenase digestion of adipose tissue. Primary adipocytes from UN offspring had significantly increased the secretion of pro-inflammatory cytokines accompanied by increased cytokine/cytokine receptor expression. This correlated with increased TLR4/NF-κB signaling. While increased inflammatory potential was not observed in adipocytes derived from UNGH offspring, there was a clear alteration in the expression of genes relating to carbohydrate and lipid metabolism along with nutrient transporters. Overall, preweaning GH treatment alters detrimental patterns of development, which predispose UN offspring to obesity and insulin resistance.

UNC5B receptor deletion exacerbates tissue injury in response to AKI.

Netrin-1 regulates cell survival and apoptosis by activation of its receptors, including UNC5B. However, the in vivo role of UNC5B in cell survival during cellular stress and tissue injury is unknown. We investigated the role of UNC5B in cell survival in response to stress using mice heterozygously expressing the UNC5B gene (UNC5B(-/flox)) and mice with targeted homozygous deletion of UNC5B in kidney epithelial cells (UNC5B(-/flox/GGT-cre)). Mice were subjected to two different models of organ injury: ischemia reperfusion injury of the kidney and cisplatin-induced nephrotoxicity. Both mouse models of UNC5B depletion had normal organ function and histology under basal conditions. After AKI, however, UNC5B(-/flox/GGT-cre) mice exhibited significantly worse renal function and damage, increased tubular apoptosis, enhanced p53 activation, and exacerbated inflammation compared with UNC5B(-/flox) and wild-type mice. shRNA-mediated suppression of UNC5B expression in cultured tubular epithelial cells exacerbated cisplatin-induced cell death in a p53-dependent manner and blunted Akt phosphorylation. Inhibition of PI3 kinase similarly exacerbated cisplatin-induced apoptosis; in contrast, overexpression of UNC5B reduced cisplatin-induced apoptosis in these cells. Taken together, these results show that the netrin-1 receptor UNC5B plays a critical role in cell survival and kidney injury through Akt-mediated inactivation of p53 in response to stress.