Identification and profiling of CXCR3–CXCR4 chemokine receptor heteromer complexes

Background and Purpose

The C-X-C chemokine receptors 3 (CXCR3) and C-X-C chemokine receptors 4 (CXCR4) are involved in various autoimmune diseases and cancers. Small antagonists have previously been shown to cross-inhibit chemokine binding to CXCR4, CC chemokine receptors 2 (CCR2) and 5 (CCR5) heteromers. We investigated whether CXCR3 and CXCR4 can form heteromeric complexes and the binding characteristics of chemokines and small ligand compounds to these chemokine receptor heteromers.

Experimental Approach

CXCR3–CXCR4 heteromers were identified in HEK293T cells using co-immunoprecipitation, time-resolved fluorescence resonance energy transfer, saturation BRET and the GPCR-heteromer identification technology (HIT) approach. Equilibrium competition binding and dissociation experiments were performed to detect negative binding cooperativity.

Key Results

We provide evidence that chemokine receptors CXCR3 and CXCR4 form heteromeric complexes in HEK293T cells. Chemokine binding was mutually exclusive on membranes co-expressing CXCR3 and CXCR4 as revealed by equilibrium competition binding and dissociation experiments. The small CXCR3 agonist VUF10661 impaired binding of CXCL12 to CXCR4, whereas small antagonists were unable to cross-inhibit chemokine binding to the other chemokine receptor. In contrast, negative binding cooperativity between CXCR3 and CXCR4 chemokines was not observed in intact cells. However, using the GPCR-HIT approach, we have evidence for specific β-arrestin2 recruitment to CXCR3-CXCR4 heteromers in response to agonist stimulation.

Conclusions and Implications

This study indicates that heteromeric CXCR3–CXCR4 complexes may act as functional units in living cells, which potentially open up novel therapeutic opportunities.     

Disparities of conjugating protective enzyme activities in the colon of patients with adenomas and carcinomas

AIM:To investigate the metabolic enzymatic capacity of the colon mucosa to detoxify noxious carcinogenic compounds.METHODS:We investigated the activity of 2 conjugating enzymes-the microsomal uridine glucuronosyltransferase(UGT)and the cytosomal glutathione S-transferase(GST)in the uninvolved mucosa of the colon transversum and sigmoideum in patients with adenomatous polyps and colorectal cancer.Biopsies were taken from the mucosa during colonoscopies which were done for clinical(diagnostic)reasons.After storage,the biopsy material was homogenized and after differential centrifugation the enzyme assays were performed with 4-nitrophenol(UGT)and 1-chloro 2,4-dinitrobenzene(GST)as substrates.RESULTS:About 48 patients were included of which28 had adenomas and 20 had colorectal carcinomas confirmed by histopathology.Enzyme activities were expressed as nmol/mg per minute protein for the GST and as pmol/mg per minute protein for the UGT.Analysis of variance(F-test)indicated that both enzymes were more widely distributed in adenoma than in cancer patients.The means±SD were smaller for cancer patients:GST for adenomas 268±152 vs 241±69 for carcinomas and UGT for adenomas 197±200 vs 150±86 for carcinomas.CONCLUSION:Compared to patients with adenomatous colon polyps those with colorectal carcinoma exhibited a lower capacity of detoxifying enzyme metabolism and their activities clustered over a smaller range.     

DNA content in malignant salivary gland tumours

Objective:  Our aim was to evaluate the DNA content in malignant salivary gland tumours using image cytometry and its possible relationships with clinical and morphologic findings, disease course and prognosis.
Patients and methods:  The study sample comprised 31 patients diagnosed and treated for primary malignant salivary gland tumours. Formalin-fixed, paraffin-embedded surgical specimens of all patients were Feulgen-stained for DNA content analysis by image cytometry. Statistical analysis was used to investigate possible relationships between DNA content variables and clinical and histological findings, disease course and patient survival.
Results:  Seventeen (55%) cases of our sample were graded as DNA diploid, four (13%) as DNA aneuploid and 10 (32%) as DNA multiploid. In 15 (48%) cases, the 5c exceeding rate (5cER) was higher than 1.7%. DNA ploidy correlated with N stage and tumour size. DNA ploidy and 5cER had a statistically significant prognostic influence on overall and disease-free survival in univariate analysis. However, in multivariate analysis, stage classification was the only parameter with an independent prognosis value.
Conclusion:  Abnormal DNA content is a common finding in salivary gland cancers. Our results suggest an important role of DNA content analysis in the evaluation of these tumours.     

Evolution of imaging for abdominal perforation

INTRODUCTION

Gastrointestinal (GI) perforation is a common surgical presentation. In recent years, computed tomography (CT) has been shown to be accurate for predicting the site of GI perforation, and has become the investigation of choice. However the signs may be subtle or only indirectly related to the site or aetiology of perforation.

SUBJECTS AND METHODS

A MEDLINE and PubMed search was performed for journals before June 2009 with MeSH major terms ‘CT’ and ‘perforation’. Non-English speaking literature was excluded.

RESULTS

Examples of GI perforation of various aetiologies are reviewed (inflammatory, neoplastic, traumatic and iatrogenic) high-lighting characteristic CT appearances as well as pitfalls in diagnosis. Features of perforation include the presence of free gas or fluid within the supra- and/or inframesocolic compartments, segmental bowel wall thickening, bowel wall discontinuity, stranding of the mesenteric fat and abscess formation.

CONCLUSIONS

These differentiating features facilitate accurate multidisciplinary pre-operative evaluation, necessary to plan patient management and potential surgical approach.     

Panton–Valentine leukocidin Staphylococcus aureus osteomyelitis of the adult tibia – a case report

Panton–Valentine leukocidin toxin producing Staphylococcus aureus (PVLSA) is known to be responsible for recurrent soft tissue infections and more serious invasive infections including necrotising pneumonia, pyomyositis, and osteomyelitis. Most reported cases involving musculoskeletal infection in adults are associated with methicillin-resistant S. aureus (MRSA) PVL-producing strains. We present the case of an adult male with PVL toxin–producing methicillin-sensitive S. aureus (MSSA) osteomyelitis of the tibia which has not previously been described in adults and highlight issues of recognition, treatment, and surgical management of PVLSA osteomyelitis.