Ultrasonographic Features of Hip Joints in Mucopolysaccharidoses Type I and II

Objectives

The primary aim of this study was to assess the ultrasonographic features of hip joints in patients with mucopolysaccharidosis (MPS) type I and II in comparison with healthy population. The secondary aims were to correlate these features with clinical measures and to evaluate the utility of ultrasound in the diagnosis of MPS disease.

Materials and Methods

Sixteen MPS I (n = 3) and II (n = 13) patients were enrolled in the present study and underwent clinical and radiological evaluation, and bilateral high-resolution ultrasonography (US) of hip joints. The distance from the femoral neck to joint capsule (synovial joint space, SJS), joint effusion, synovial hyperthrophy, and local pathological vascularization were evaluated. The results were compared to the healthy population and correlated with clinical and radiological measures.

Results

1. There was a difference in US SJS between children with MPS disease and the normative value for healthy population (7mm). Mean values of SJS were 15.81 ± 4.08 cm (right hip joints) and 15.69 ± 4.19 cm (left joints). 2. No inflammatory joint abnormalities were detected in MPS patients. 3. There was a clear correlation between US SJS and patients’ age and height, while no clear correlation was observed between SJS and disease severity.

Conclusions

1. Patients with MPS I and II present specific features in hip joint ultrasonography. 2. The data suggests that ultrasonography might be effective in the evaluation of hip joint involvement in patients with MPS and might present a valuable tool in facilitating the diagnosis and follow up of the disease.     

Expression Profile of Cytokines and Enzymes mRNA in Blood Leukocytes of Dogs with Leptospirosis and Its Associated Pulmonary Hemorrhage Syndrome

Background

Dogs with leptospirosis show similar organ manifestations and disease course as human patients, including acute kidney injury and pulmonary hemorrhage, making this naturally-occurring infection a good animal model for human leptospirosis. Expression patterns of cytokines and enzymes have been correlated with disease manifestations and clinical outcome in humans and animals. The aim of this study was to describe mRNA expression of pro- and anti-inflammatory mediators in canine leptospirosis and to compare it with other renal diseases to identify patterns characterizing the disease and especially its pulmonary form.

Methodology and Principal Findings

The mRNA abundance of cytokines (IL-1α, IL-1β, IL-8, IL-10, TNF-α, TGF-β) and enzymes (5-LO, iNOS) was measured prospectively in blood leukocytes from 34 dogs with severe leptospirosis and acute kidney injury, including 22 dogs with leptospirosis-associated pulmonary hemorrhages. Dogs with leptospirosis were compared to 14 dogs with acute kidney injury of other origin than leptospirosis, 8 dogs with chronic kidney disease, and 10 healthy control dogs. Canine leptospirosis was characterized by high 5-LO and low TNF-α expression compared to other causes of acute kidney injury, although the decreased TNF-α expression was also seen in chronic kidney disease. Leptospirosis-associated pulmonary hemorrhage was not characterized by a specific pattern, with only mild changes noted, including increased IL-10 and decreased 5-LO expression on some days in affected dogs. Fatal outcome from pulmonary hemorrhages was associated with low TNF-α, high IL-1β, and high iNOS expression, a pattern possibly expressed also in dogs with other forms of acute kidney injury.

Conclusion

The patterns of cytokine and enzyme expression observed in the present study indicate a complex pro- and anti-inflammatory response to the infection with leptospires. The recognition of these signatures may be of diagnostic and prognostic relevance for affected individuals and they may indicate options for newer therapies targeting the identified pathways.     

Decreased Histone Deacetylase 2 (HDAC2) in Peripheral Blood Monocytes (PBMCs) of COPD Patients

Background

Histone deacetylase 2 (HDAC2) is a class I histone deacetylase family member that plays a critical role in suppressing inflammatory gene expression in the airways, lung parenchyma, and alveolar macrophages in patients with chronic obstructive pulmonary disease (COPD). However, the expression of HDAC2 in peripheral blood monocytes (PBMCs), nuclear factor kappa B (NF-κB) p65, and serum inflammatory cytokine levels in COPD patients, smokers, and non-smokers remains unclear.

Methods

PBMCs were obtained from COPD patients, healthy smokers, and healthy nonsmokers. The HDAC2 and NF-κB p65 expression were quantified by Western Blot. HDAC activity was assessed by an HDAC fluorometric immunoprecipitation activity assay kit. Serum tumor necrosis factor-alpha (TNF-α) and interleukin-8 (IL-8) levels were measured by ELISA.

Results

HDAC2 expression and HDAC activity were decreased in PBMCs in COPD patients compared with smokers and non-smokers. Increased NF-κB p65 expression, serum TNF-α and IL-8 levels were observed in COPD patients compared with nonsmokers. The FEV₁%pred was positively correlated with HDAC2 expression and HDAC activity in COPD patients. Smokers had decreased HDAC activity, increased NF-κB p65 expression and serum TNF-α compared with nonsmokers.

Conclusions

HDAC2 expression was decreased in PBMCs of COPD patients and was correlated with disease severity. The reduction of HDAC2 expression not only directly enhances the expression of inflammatory genes, but may account for the activation of NF-κB mediated inflammation. Decreased HDAC2 may serve as a potential biomarker of COPD and predict the decline of lung function.     

Pathway Implications of Aberrant Global Methylation in Adrenocortical Cancer

Context

Adrenocortical carcinomas (ACC) are a rare tumor type with a poor five-year survival rate and limited treatment options.

Objective

Understanding of the molecular pathogenesis of this disease has been aided by genomic analyses highlighting alterations in TP53, WNT, and IGF signaling pathways. Further elucidation is needed to reveal therapeutically actionable targets in ACC.

Design

In this study, global DNA methylation levels were assessed by the Infinium HumanMethylation450 BeadChip Array on 18 ACC tumors and 6 normal adrenal tissues. A new, non-linear correlation approach, the discretization method, assessed the relationship between DNA methylation/gene expression across ACC tumors.

Results

This correlation analysis revealed epigenetic regulation of genes known to modulate TP53, WNT, and IGF signaling, as well as silencing of the tumor suppressor MARCKS, previously unreported in ACC.

Conclusions

DNA methylation may regulate genes known to play a role in ACC pathogenesis as well as known tumor suppressors.     

Naringin Reduces Hyperglycemia-Induced Cardiac Fibrosis by Relieving Oxidative Stress

Introduction

Hyperglycemia promotes myocardial fibrotic lesions through upregulation of PKC and p38 in response to redox changes. The effects of naringin on hyperglycemia-induced myocardial fibrotic changes and its putative effects on PKC-β and p38 protein expression in type 1 rat model of diabetes are hereby investigated.

Methods

Male Sprague-Dawley rats were divided into six groups I-VI. Groups I and II, were orally treated with distilled water {3.0 ml/kg body weight (BW)} and naringin (50 mg/kg BW), respectively. Groups III, IV, V and VI were rendered diabetic by a single intraperitoneal injection of streptozotocin (60 mg/kg, BW) and were similarly treated with subcutaneous insulin (8.0 I.U/kg BW, twice daily), naringin (50 mg/kg BW), distilled water (3.0 ml/Kg BW) and ramipril (3.0 mg/kg/BW), respectively. The animals were sacrificed after 56 days by halothane overdose; blood and heart samples removed for further analysis.

Results

The untreated diabetic rats exhibited significantly increased oxidative stress, NADPH oxidase activity, increased cardiac fibrosis, PKC-β and p38 mitogen activated protein kinase expression compared to controls. Naringin treatment significantly ameliorated these changes in diabetic rats compared to the untreated diabetic controls.

Conclusions

Naringin’s amelioration of myocardial fibrosis by modulating p38 and PKC-β protein expression possibly through its known antioxidant actions and may therefore be useful in retarding the progression of fibrosis in a diabetic heart.     

Sequestration of Vascular Endothelial Growth Factor (VEGF) Induces Late Restrictive Lung Disease

Rationale

Neonatal respiratory distress syndrome is a restrictive lung disease characterized by surfactant deficiency. Decreased vascular endothelial growth factor (VEGF), which demonstrates important roles in angiogenesis and vasculogenesis, has been implicated in the pathogenesis of restrictive lung diseases. Current animal models investigating VEGF in the etiology and outcomes of RDS require premature delivery, hypoxia, anatomically or temporally limited inhibition, or other supplemental interventions. Consequently, little is known about the isolated effects of chronic VEGF inhibition, started at birth, on subsequent developing lung structure and function.

Objectives

To determine whether inducible, mesenchyme-specific VEGF inhibition in the neonatal mouse lung results in long-term modulation of AECII and whole lung function.

Methods

Triple transgenic mice expressing the soluble VEGF receptor sFlt-1 specifically in the mesenchyme (Dermo-1/rtTA/sFlt-1) were generated and compared to littermate controls at 3 months to determine the impact of neonatal downregulation of mesenchymal VEGF expression on lung structure, cell composition and function. Reduced tissue VEGF bioavailability has previously been demonstrated with this model.

Measurements and Main Results

Triple transgenic mice demonstrated restrictive lung pathology. No differences in gross vascular development or protein levels of vascular endothelial markers was noted, but there was a significant decrease in perivascular smooth muscle and type I collagen. Mutants had decreased expression levels of surfactant protein C and hypoxia inducible factor 1-alpha without a difference in number of type II pneumocytes.

Conclusions

These data show that mesenchyme-specific inhibition of VEGF in neonatal mice results in late restrictive disease, making this transgenic mouse a novel model for future investigations on the consequences of neonatal RDS and potential interventions.     

Suppressive Role of PPARγ-Regulated Endothelial Nitric Oxide Synthase in Adipocyte Lipolysis

Introduction

Metabolic syndrome causes insulin resistance and is associated with risk factor clustering, thereby increasing the risk of atherosclerosis. Recently, endothelial nitric oxide synthase deficient (eNOS-/-) mice have been reported to show metabolic disorders. Interestingly, eNOS has also been reported to be expressed in non-endothelial cells including adipocytes, but the functions of eNOS in adipocytes remain unclear.

Methods and Results

The eNOS expression was induced with adipocyte differentiation and inhibition of eNOS/NO enhanced lipolysis in vitro and in vivo. Furthermore, the administration of a high fat diet (HFD) was able to induce non-alcoholic steatohepatitis (NASH) in eNOS-/- mice but not in wild type mice. A PPARγ antagonist increased eNOS expression in adipocytes and suppressed HFD-induced fatty liver changes.

Conclusions

eNOS-/- mice induce NASH development, and these findings provide new insights into the therapeutic approach for fatty liver disease and related disorders.     

Mefunidone Attenuates Tubulointerstitial Fibrosis in a Rat Model of Unilateral Ureteral Obstruction

Background

Inflammation has a crucial role in renal interstitial fibrosis, which is the common pathway of chronic kidney diseases. Mefunidone (MFD) is a new compound which could effectively inhibit the proliferation of renal fibroblasts in vitro. However, the overall effect of Mefunidone in renal fibrosis remains unknown.

Methods

Sprague-Dawley rats were randomly divided intro 6 groups: sham operation, unilateral ureteral obstruction (UUO), UUO/Mefunidone (25, 50, 100mg/kg/day) and UUO/PFD (500mg/kg/day). The rats were sacrificed respectively on days 3, 7, and 14 after the operation. Tubulointerstitial injury index, interstitial collagen deposition, expression of fibronectin (FN), α-smooth muscle actin (α-SMA), type I and III collagen and the number of CD3+ and CD68+ cells were determined. The expressions of proinflammatory cytokines, p-ERK, p-IκB, and p-STAT3 were measured in human renal proximal tubular epithelial cells of HK-2 or macrophages.

Results

Mefunidone treatment significantly attenuated tubulointerstitial injury, interstitial collagen deposition, expression of FN, α-SMA, type I and III collagen in the obstructive kidneys, which correlated with significantly reduced the number of T cells and macrophages in the obstructive kidneys. Mechanistically, Mefunidone significantly inhibited tumor necrosis factor-α (TNF-α-) or lipopolysaccharide (LPS)-induced production of proinflammatory cytokines. This effect is possibly due to the inhibition of phosphorylation of ERK, IκB, and STAT3.

Conclusion

Mefunidone treatment attenuated tubulointerstitial fibrosis in a rat model of UUO, at least in part, through inhibition of inflammation.     

Loss of Endothelial Barrier in Marfan Mice (mgR/mgR) Results in Severe Inflammation after Adenoviral Gene Therapy

Objectives

Marfan syndrome is an autosomal dominant inherited disorder of connective tissue. The vascular complications of Marfan syndrome have the biggest impact on life expectancy. The aorta of Marfan patients reveals degradation of elastin layers caused by increased proteolytic activity of matrix metalloproteinases (MMPs). In this study we performed adenoviral gene transfer of human tissue inhibitor of matrix metalloproteinases-1 (hTIMP-1) in aortic grafts of fibrillin-1 deficient Marfan mice (mgR/mgR) in order to reduce elastolysis.

Methods

We performed heterotopic infrarenal transplantation of the thoracic aorta in female mice (n = 7 per group). Before implantation, mgR/mgR and wild-type aortas (WT, C57BL/6) were transduced ex vivo with an adenoviral vector coding for human TIMP-1 (Ad.hTIMP-1) or β-galactosidase (Ad.β-Gal). As control mgR/mgR and wild-type aortas received no gene therapy. Thirty days after surgery, overexpression of the transgene was assessed by immunohistochemistry (IHC) and collagen in situ zymography. Histologic staining was performed to investigate inflammation, the neointimal index (NI), and elastin breaks. Endothelial barrier function of native not virus-exposed aortas was evaluated by perfusion of fluorescent albumin and examinations of virus-exposed tissue were performed by transmission electron microscopy (TEM).

Results

IHC and ISZ revealed sufficient expression of the transgene. Severe cellular inflammation and intima hyperplasia were seen only in adenovirus treated mgR/mgR aortas (Ad.β-Gal, Ad.hTIMP-1 NI: 0.23; 0.43), but not in native and Ad.hTIMP-1 treated WT (NI: 0.01; 0.00). Compared to native mgR/mgR and Ad.hTIMP-1 treated WT aorta, the NI is highly significant greater in Ad.hTIMP-1 transduced mgR/mgR aorta (p = 0.001; p = 0.001). As expected, untreated Marfan grafts showed significant more elastolysis compared to WT (p = 0.001). However, elastolysis in Marfan aortas was not reduced by adenoviral overexpression of hTIMP-1 (compared to untreated Marfan aorta: Ad.hTIMP-1 p = 0.902; control Ad.β-Gal. p = 0.165). The virus-untreated and not transplanted mgR/mgR aorta revealed a significant increase of albumin diffusion through the endothelial barrier (p = 0.037). TEM analysis of adenovirus-exposed mgR/mgR aortas displayed disruption of the basement membrane and basolateral space.

Conclusions

Murine Marfan aortic grafts developed severe inflammation after adenoviral contact. We demonstrated that fibrillin-1 deficiency is associated with relevant dysfunction of the endothelial barrier that enables adenovirus to induce vessel-harming inflammation. Endothelial dysfunction may play a pivotal role in the development of the vascular phenotype of Marfan syndrome.     

Analysis of the Proteolytic Processing of ABCA3: Identification of Cleavage Site and Involved Proteases

Rationale

ABCA3 is a lipid transporter in the limiting membrane of lamellar bodies in alveolar type II cells. Mutations in the ABCA3 gene cause respiratory distress syndrome in new-borns and childhood interstitial lung disease. ABCA3 is N-terminally cleaved by an as yet unknown protease, a process believed to regulate ABCA3 activity.

Methods

The exact site where ABCA3 is cleaved was localized using mass spectrometry (MS). Proteases involved in ABCA3 processing were identified using small molecule inhibitors and siRNA mediated gene knockdown. Results were verified by in vitro digestion of a synthetic peptide substrate mimicking ABCA3’s cleavage region, followed by MS analysis.

Results

We found that cleavage of ABCA3 occurs after Lys¹⁷⁴ which is located in the proteins’ first luminal loop. Inhibition of cathepsin L and, to a lesser extent, cathepsin B resulted in attenuation of ABCA3 cleavage. Both enzymes showed activity against the ABCA3 peptide in vitro with cathepsin L being more active.

Conclusion

We show here that, like some other proteins of the lysosomal membrane, ABCA3 is a substrate of cathepsin L. Therefore, cathepsin L may represent a potential target to therapeutically influence ABCA3 activity in ABCA3-associated lung disease.     

Airway Surface Dehydration Aggravates Cigarette Smoke-Induced Hallmarks of COPD in Mice

Introduction

Airway surface dehydration, caused by an imbalance between secretion and absorption of ions and fluid across the epithelium and/or increased epithelial mucin secretion, impairs mucociliary clearance. Recent evidence suggests that this mechanism may be implicated in chronic obstructive pulmonary disease (COPD). However, the role of airway surface dehydration in the pathogenesis of cigarette smoke (CS)-induced COPD remains unknown.

Objective

We aimed to investigate in vivo the effect of airway surface dehydration on several CS-induced hallmarks of COPD in mice with airway-specific overexpression of the β-subunit of the epithelial Na⁺ channel (βENaC).

Methods

βENaC-Tg mice and wild-type (WT) littermates were exposed to air or CS for 4 or 8 weeks. Pathological hallmarks of COPD, including goblet cell metaplasia, mucin expression, pulmonary inflammation, lymphoid follicles, emphysema and airway wall remodelling were determined and lung function was measured.

Results

Airway surface dehydration in βENaC-Tg mice aggravated CS-induced airway inflammation, mucin expression and destruction of alveolar walls and accelerated the formation of pulmonary lymphoid follicles. Moreover, lung function measurements demonstrated an increased compliance and total lung capacity and a lower resistance and hysteresis in βENaC-Tg mice, compared to WT mice. CS exposure further altered lung function measurements.

Conclusions

We conclude that airway surface dehydration is a risk factor that aggravates CS-induced hallmarks of COPD.     

No Fabry Disease in Patients Presenting with Isolated Small Fiber Neuropathy

Objective

Screening for Fabry disease in patients with small fiber neuropathy has been suggested, especially since Fabry disease is potentially treatable. However, the diagnostic yield of testing for Fabry disease in isolated small fiber neuropathy patients has never been systematically investigated. Our aim is to determine the presence of Fabry disease in patients with small fiber neuropathy.

Methods

Patients referred to our institute, who met the criteria for isolated small fiber neuropathy were tested for Fabry disease by measurement of alpha-Galactosidase A activity in blood, lysosomal globotriaosylsphingosine in urine and analysis on possible GLA gene mutations.

Results

725 patients diagnosed with small fiber neuropathy were screened for Fabry disease. No skin abnormalities were seen except for redness of the hands or feet in 30.9% of the patients. Alfa-Galactosidase A activity was tested in all 725 patients and showed diminished activity in eight patients. Lysosomal globotriaosylsphingosine was examined in 509 patients and was normal in all tested individuals. Screening of GLA for mutations was performed for 440 patients, including those with diminished α-Galactosidase A activity. Thirteen patients showed a GLA gene variant. One likely pathogenic variant was found in a female patient. The diagnosis Fabry disease could not be confirmed over time in this patient. Eventually none of the patients were diagnosed with Fabry disease.

Conclusions

In patients with isolated small fiber neuropathy, and no other signs compatible with Fabry disease, the diagnostic yield of testing for Fabry disease is extremely low. Testing for Fabry disease should be considered only in cases with additional characteristics, such as childhood onset, cardiovascular disease, renal failure, or typical skin lesions.     

Elevated Levels of IFN-γ in CSF and Serum of Patients with Amyotrophic Lateral Sclerosis

Objectives

To explore whether the levels of IFN-γ in cerebral spinal fluid (CSF) and serum are elevated in ALS patients and to analyze the correlations between the IFN-γ levels and disease progression.

Methods

CSF and serum samples were obtained from 52 ALS patients and 31 non-ALS patients. The levels of IFN-γ in CSF and serum were assessed, and disease progression parameters, including the disease interval (months from onset, MFO), the revised ALS Functional Rating Scale (ALSFRS-r) score and the disease progression rate (DPR) were analyzed by registered neurologists. All samples were measured using a commercial enzyme-linked immunosorbent assay. Statistical analyses were performed using Prism software.

Results

Compared to the non-ALS patients, the ALS patients displayed significantly increased levels of IFN-γ in both CSF and serum, and these values consistently correlated with disease progression.

Conclusions

These results demonstrated that IFN-γ in CSF may serve as a biomarker of ALS differentiation and progression. CSF IFN-γ was a more reliable biomarker of disease diagnosis and progression than serum IFN-γ.     

Examination of a Viral Infection Mimetic Model in Human iPS Cell-Derived Insulin-Producing Cells and the Anti-Apoptotic Effect of GLP-1 Analogue

Aims

Viral infection is associated with pancreatic beta cell destruction in fulminant type 1 diabetes mellitus. The aim of this study was to investigate the acceleration and protective mechanisms of beta cell destruction by establishing a model of viral infection in pancreatic beta cells.

Methods

Polyinosinic:polycytidylic acid was transfected into MIN6 cells and insulin-producing cells differentiated from human induced pluripotent stem cells via small molecule applications. Gene expression was analyzed by real-time PCR, and apoptosis was evaluated by caspase-3 activity and TUNEL staining. The anti-apoptotic effect of Exendin-4 was also evaluated.

Results

Polyinosinic:polycytidylic acid transfection led to elevated expression of the genes encoding IFNα, IFNβ, CXCL10, Fas, viral receptors, and IFN-inducible antiviral effectors in MIN6 cells. Exendin-4 treatment suppressed the elevated gene expression levels and reduced polyinosinic:polycytidylic acid-induced apoptosis both in MIN6 cells and in insulin-producing cells from human induced pluripotent stem cells. Glucagon-like peptide-1 receptor, protein kinase A, and phosphatidylinositol-3 kinase inhibitors counteracted the anti-apoptotic effect of Exendin-4.

Conclusions

Polyinosinic:polycytidylic acid transfection can mimic viral infection, and Exendin-4 exerted an anti-apoptotic effect both in MIN6 and insulin-producing cells from human induced pluripotent stem cells.     

Positron Emission Tomography and Magnetic Resonance Imaging of the Brain in Fabry Disease: A Nationwide,Long-Time,Prospective Follow-Up

Background

Fabry disease is a rare metabolic glycosphingolipid storage disease caused by deficiency of the lysosomal enzyme α-galactosidase A—leading to cellular accumulation of globotriasylceramide in different organs, vessels, tissues, and nerves. The disease is associated with an increased risk of cerebrovascular disease at a young age in addition to heart and kidney failure.

Objective

The objective of this study was to assess brain function and structure in the Danish cohort of patients with Fabry disease in a prospective way using 18-fluoro-deoxyglucose (F-18 FDG) positron emission tomography (PET) and magnetic resonance imaging (MRI).

Patients

Forty patients with Fabry disease (14 males, 26 females, age at inclusion: 10–66 years, median: 39 years) underwent a brain F-18-FDG-PET-scan at inclusion, and 31 patients were followed with FDG-PET biannually for up to seven years. All patients (except one) had a brain MRI-scan at inclusion, and 34 patients were followed with MRI biannually for up to nine years.

Image Analysis

The FDG-PET-images were inspected visually and analysed using a quantitative 3-dimensional stereotactic surface projection analysis (Neurostat). MRI images were also inspected visually and severity of white matter lesions (WMLs) was graded using a visual rating scale.

Results

In 28 patients brain-FDG-PET was normal; in 23 of these 28 patients brain MRI was normal—of the remaining five patients in this group, four patients had WMLs and one patient never had an MRI-scan. In 10 patients hypometabolic areas were observed on brain-FDG-PET; all of these patients had cerebral infarcts/hemorrhages visualized on MRI corresponding to the main hypometabolic areas. In two patients brain-FDG-PET was ambiguous, while MRI was normal in one and abnormal in the other.

Conclusion

Our data indicated that, in patients with Fabry disease, MRI is the preferable clinical modality—if applicable—when monitoring cerebral status, as no additional major brain-pathology was detected with FDG-PET.     

The Leukocyte Immunoglobulin-Like Receptor Family Member LILRB5 Binds to HLA-Class I Heavy Chains

Objective

The leukocyte immunoglobulin-like receptor (LILR) family includes inhibitory and stimulatory members which bind to classical and non-classical HLA-class I. The ligands for many LILR including LILRB5 have not yet been identified.

Methods

We generated C-terminal eGFP and N-terminal FLAG-tagged fusion constructs for monitoring LILR expression. We screened for LILR binding to HLA-class I by tetramer staining of 293T cells transfected with LILRA1, A4, A5 A6 and LILRB2 and LILRB5. We also studied HLA class I tetramer binding to LILRB5 on peripheral monocyte cells. LILRB5 binding to HLA-class I heavy chains was confirmed by co-immunoprecipitation.

Results

HLA-B27 (B27) free heavy chain (FHC) dimer but not other HLA-class I stained LILRB5-transfected 293T cells. B27 dimer binding to LILRB5 was blocked with the class I heavy chain antibody HC10 and anti-LILRB5 antisera. B27 dimers also bound to LILRB5 on peripheral monocytes. HLA-B7 and B27 heavy chains co-immunoprecipitated with LILRB5 in transduced B and rat basophil RBL cell lines.

Conclusions

Our findings show that class I free heavy chains are ligands for LILRB5. The unique binding specificity of LILRB5 for HLA-class I heavy chains probably results from differences in the D1 and D2 immunoglobulin-like binding domains which are distinct from other LILR which bind to β2m-associated HLA-class I.     

Dystroglycan Depletion Impairs Actin-Dependent Functions of Differentiated Kasumi-1 Cells

Background

Dystroglycan has recently been characterised in blood tissue cells, as part of the dystrophin glycoprotein complex involved in the differentiation process of neutrophils.

Purpose

In the present study we have investigated the role of dystroglycan in the human promyelocytic leukemic cell line Kasumi-1 differentiated to macrophage-like cells.

Methods

We characterised the pattern expression and subcellular distribution of dystroglycans in non-differentiated and differentiated Kasumi-1 cells.

Results

Our results demonstrated by WB and flow cytometer assays that during the differentiation process to macrophages, dystroglycans were down-regulated; these results were confirmed with qRT-PCR assays. Additionally, depletion of dystroglycan by RNAi resulted in altered morphology and reduced properties of differentiated Kasumi-1 cells, including morphology, migration and phagocytic activities although secretion of IL-1β and expression of markers of differentiation are not altered.

Conclusion

Our findings strongly implicate dystroglycan as a key membrane adhesion protein involved in actin-based structures during the differentiation process in Kasumi-1 cells.     

Therapeutic Effects of Tangshen Formula on Diabetic Nephropathy in Rats

Objective

Inflammation and fibrosis are essential promoters in the pathogenesis of diabetic nephropathy (DN) in type 2 diabetes. The present study examined the anti-inflammation and anti-fibrosis effect of Tangshen Formula (TSF), a traditional Chinese medicine, on DN.

Research Design and Methods

Protective role of TSF in DN was examined in a rat model of type 2 DN that was established by high-fat diet-fed and low-dose-streptozotocin injection. TSF was suspended in 0.5% CMC-Na solution and delivered by oral gavage at a dosage of 1.67g/Kg body weight/day. The therapeutic effects and mechanisms of TSF on diabetic kidney injury were examined.

Results

We found that TSF treatment for 20 weeks attenuated DN by significantly inhibiting urinary excretion of albumin and renal histological injuries. These beneficial effects were associated with an inactivation of NF-κB signaling, thereby blocking the upregulation of pro-inflammatory cytokines (IL-1β, TNFα), chemokine (MCP-1), and macrophage infiltration in the TSF-treated rats with type 2 DN. In addition, TSF treatment also inactivated TGF-β/Smad3 signaling and therefore suppressed renal fibrosis including expressions of fibronectin, collagen I, and collagen IV. Further studies revealed that the inhibitory effect of TSF on TGF-β/Smad3 and NF-κB signaling in DN was associated with inhibition of Smurf2-dependent ubiquitin degradation of Smad7.

Conclusions

The present study reveals that TSF has therapeutic potential for type 2 DN in rats. Blockade of NF-κB-driven renal inflammation and TGF-β/Smad3-mediated renal fibrosis by preventing the Smurf2-mediated Smad7 degradation pathway may be mechanisms through which TSF inhibits type 2 DN.