C-type lectin protein isoforms of Macrovipera lebetina: cDNA cloning and genetic diversity

Snake venom contains a complex protein mixture belonging to a few well-characterized protein families: disintegrins, phospholipase A2, serine protease, l-amino acid oxidase, Zn-dependent metalloproteinase, natriuretic peptides, myotoxins, cysteine-rich secretory protein (CRISP) toxins, Kunitz-type protease inhibitors and C-type lectin-like. Despite their pharmacological importance, little is known about the exact composition of each protein family. We report here the cloning of 25 complete ORFs from Macrovipera lebetina transmediterranea venom gland that encodes several isoforms and novel C-type lectins (CTLs). 16 alpha and nine beta CTL chains were identified. Based on their sequence alignment, we categorized the 16 CTL alpha subunits into five groups and the nine CTL beta subunits into four groups to deduce the phylogenetic tree of M. lebetina transmediterranea CTLs. Sequence analysis revealed that they share a high degree of similarity with each other and with other snake venom CTLs. The M. lebetina transmediterranea CTL sequences described here contain a C-lectin carbohydrate recognition domain-like fold (C-lectin CRD-like) characterized by several conserved amino acid residues in their structure, especially the cysteine. Finally, based on the comparison of some Macrovipera CTL, we propose that some new CTL gene versions should have occurred through “domains shuffling” from former genes.     

Acute pseudoseptic arthritis after viscosupplementation of the knee: a case report

Intra-articular injections of hyaluronan are generally well tolerated; the most common adverse event associated with their use is an inflammatory reaction or a flare at the injection site. Naturally derived sodium hyaluronates have not been associated with pseudosepsis; we describe a case of severe arthritis mimicking acute septic arthritis after naturally hyaluronan. A 70-year-old woman had a medial femorotibial and patellofemoral compartment knee osteoarthritis. Hyaluronic acid injection was indicated because of the persistence of a pain while walking and restricted ability to walk 100 m, as well as the installation of a flessum. She was admitted for an evaluation of a joint effusion occurring within 48 h after the first hyaluronic acid intra-articular injection (first course), with no fever or chills. Joint fluid was found to be cloudy and contained 24,000 cells per mm(3). A knee aspiration was performed to evaluate the possibility of a joint infection. The examination of synovial fluid under phase contrast and polarizing microscopes showed no crystals, and culture on standard media was negative. Acute septic arthritis was considered to be the most likely diagnosis, and probabilistic antibiotic therapy was started. The result of the bacteriological examinations also returned negative. The absence of general signs, particularly fever and chills, with fast effusion regression in less than 4 days did not plead in favor of a septic origin. In front of this beam of arguments, antibiotic therapy was stopped after 4 days. The nonsteroidal anti-inflammatory drug (Nimésulide) was given. The clinical and laboratory test abnormalities were normalized within 2 weeks. Our case has the particularity to occur after using Curavisc, which is known as producing no allergenic reactions. Moreover, the described reaction occurred with the very first infiltration within the very first course.     

Hyperparathyroidism-jaw tumor syndrome: A case report

We report the clinical and genetic findings in a 23-year-old woman with hyperparathyroidism-jaw tumor syndrome (HPT-JT). The patient had a family history of primary hyperparathyroidism (PHPT) and uterine fibroma in her mother. The patient presented muscle weakness. The diagnosis of PHPT was confirmed by an elevated parathyroid hormone level above 1450 pg/ml with hypercalcemia and hypercalciuria. X-ray radiographies showed a radiolucent lesion in the right body of the mandible. Bilateral neck exploration was performed. An inferior right parathyroidectomy, a left thyroid lobectomy with isthmectomy and thymectomy were carried out. Histopathological examination of the specimen showed a diffuse hyperplasia of the parathyroid principal cells. The association of PHPT with a right jaw tumor and uterine fibroma suggested the diagnosis of HPT-JT syndrome. Mutation screening of HRPT2 gene was carried out and identified a germline mutation, consisting in a base deletion in exon 1, 85delG, inducing a frameshift. The diagnosis of HPT-JT syndrome is clinically important because of its hereditary component and its high risk of parathyroid malignancy, making a genetic inquiry necessary.     

Modulation of cellular redox state underlies antagonism between oxaliplatin and cetuximab in human colorectal cancer cell lines

Background and purpose:

Oxaliplatin is the first platinum-based compound effective in the treatment of colorectal cancer. Oxaliplatin combined with cetuximab for metastatic colorectal cancer is under evaluation. The preliminary results seem controversial, particularly for the use of cetuximab in K-Ras mutated patients. K-Ras mutation is known to affect redox homeostasis. Here we evaluated how the efficacy of oxaliplatin alone or combined with cetuximab varied according to the Ras mutation and redox status in a panel of colorectal tumour cell lines.

Experimental approach:

Viability was evaluated by methylthiazoletetrazolium assay, reactive oxygen species production by DCFDA and lucigenin on HT29-D4, Caco-2, SW480 and SW620 cell lines.

Key results:

Combination of oxaliplatin and cetuximab was less cytotoxic than oxaliplatin alone in colorectal cells harbouring wild-type Ras and membrane expression of receptors for epidermal growth factor receptor (EGFR), such as HT29-D4 and Caco-2 cells. In contrast, cetuximab did not affect oxaliplatin efficiency in cells harbouring K-Ras^V12 mutation, irrespective of membrane EGFR expression (SW620 and SW480 cells). Transfection of HT29-D4 with K-Ras^V12 decreased oxaliplatin IC₅₀ and impaired cetuximab sensitivity, without affecting expression of membrane EGFR compared with HT29-D4 control. Oxaliplatin efficacy relies on endogenous production of H₂O₂. Cetuximab inhibits H₂O₂ production inhibiting the EGFR/Nox1 NADPH oxidase pathway. Oxaliplatin efficacy was impaired by short hairpin RNA for Nox1 and by catalase (H₂O₂ scavenger).

Conclusions and implications:

Cetuximab limited oxaliplatin efficiency by affecting the redox status of cancer cells through Nox1. Such combined therapy might be improved by controlling H₂O₂ elimination.     

Effects of urokinase-type plasminogen activator in the acquisition, expression and reinstatement of cocaine-induced conditioned-place preference

Cocaine and many other psychostimulants strongly induce urokinase-type plasminogen activator (uPA) expression in the mesolimbic dopaminergic pathway, which plays a major role in drug-mediated behavioral plasticity [Bahi A, Boyer F, Gumy C, Kafri T, Dreyer JL. In vivo gene delivery of urokinase-type plasminogen activator with regulatable lentivirus induces behavioral changes in chronic cocaine administration. Eur J Neurosci 2004;20:3473-88; Bahi A, Boyer F, Kafri T, Dreyer JL. Silencing urokinase in the ventral tegmental area in vivo induces changes in cocaine-induced hyperlocomotion. J Neurochem 2006;98:1619-31; Bahi A, Dreyer JL. Overexpression of plasminogen activators in the nucleus accumbens enhances cocaine-, amphetamine- and morphine-induced reward and behavioral sensitization. Genes Brain Behav 2007]. In this study, the role of mesolimbic dopamine (DA) pathways in the development of cocaine reward was examined by conditioned-place preference in rats with bilateral intra-accumbens injections of uPA-expressing lentiviral vectors. We show that overexpression of uPA in the Nac significantly augments cocaine-induced place preference. Furthermore, while this did not affect the ability of preference to be extinguished, reinstatement with a low dose of cocaine produced significantly greater preference to the cocaine-associated context. Once CPP had been established, and the preference extinguished, reinstatement induced by a priming dose of cocaine was facilitated by uPA. Inhibition of this serine protease expression using doxycycline abolished the augmented acquisition produced by overexpression of uPA but not the expression of the cocaine-induced CPP. When uPA is inhibited during the acquisition phase, animals no longer demonstrate place preference for the environment previously paired with cocaine. B428, a specific uPA inhibitor does not affect drug reinstatement after extinction if uPA has been activated during acquisition, a clear indication that uPA is involved in the acquisition phase of conditioned-place preference. Our results suggest that that increased uPA expression with repeated drug exposure produces conditions for enhanced acquisition of cocaine-induced CPP, indicating that cocaine-induced CPP and reinstatement may be dependent on active extracellular uPA.     

A comparison of the performance of microsatellite and methylation urine analysis for predicting the recurrence of urothelial cell carcinoma, and definition of a set of markers by Bayesian network analysis

OBJECTIVE

To compare the potential of two diagnostic methods for detecting recurrence of urothelial cell carcinoma (UCC) of the bladder, by (i) detecting alterations in microsatellite DNA markers and loss of heterozygosity (LOH), and (ii) detecting aberrant gene hypermethylation, as UCC has a high recurrence rate in the urinary tract and the disease can invade muscle if new tumours are overlooked.

PATIENTS AND METHODS

Over 1 year, urine samples were retrieved from 40 patients already diagnosed with bladder UCC (30 pTa, two pTis, eight pT1). Samples were collected 6 months after bladder tumour resection, during the follow‐up schedule. We used samples to analyse nine microsatellite markers and the methylation status of 11 gene promoters. Receiver operating characteristic curves were generated and Bayesian statistics used to create an interaction network between recurrence and the biomarkers.

RESULTS

During the study, 15 of the 40 patients (38%) had a tumour recurrence and 14 were identified by cystoscopy (reference method). Overall, microsatellite markers (area under curve, AUC 0.819, 95% confidence interval, CI, 0.677–0.961) had better performance characteristics than promoter hypermethylation (AUC 0.448, 0.259–0.637) for detecting recurrence. A marker panel of IFNA, MBP, ACTBP2, D9S162 and of RASSF1A, and WIF1 generated a higher diagnostic accuracy of 86% (AUC 0.92, 0.772–0.981).

CONCLUSION

Microsatellite markers have better performance characteristics than promoter hypermethylation for detecting UCC recurrence. These data support the further development of a combination of only six markers from both methods in urinary DNA. Once validated, it could be used routinely during the follow‐up for the early detection and surveillance of UCC from the lower and upper urinary tract.     

Clinical presentation and outcome of bladder schistosoma-unrelated squamous cell carcinoma: report on 33 consecutive cases

BACKGROUND: Pure squamous cell carcinoma (SCC) is a rare entity in Western countries. The management of SCC still remains similar to that of transitional carcinoma, although it is a different entity. A retrospective review is helpful in understanding the epidemiologic aspects, pathogenesis, and treatment and prognosis of schistosoma-unrelated SCC. PATIENTS AND METHODS: Between 1987 and 2002, 30 men and 3 women had been treated for pure SCC of the bladder, not related to bilharzias. RESULTS: The mean age of patients was 59 years (range, 29-83 years). They constitute 1.2% of all bladder tumors. At presentation, 2 patients had a distant metastasis. Eleven patients (34%) had a bladder stone with recurrent urinary tract infection; they had been treated since age 6-17 years. Twenty-one patients underwent radical cystectomy, followed by chemotherapy in 2 cases. At a mean follow-up of 5 years, 14 patients (66.6%) died of locoregional disease, with associated metastasis in 5 cases. Seven patients (33.3%) are alive without any evidence of disease. The survival rate was correlated with tumor stage. CONCLUSION: The unrelated SCC has an unfavorable prognosis, mostly caused by the locally advanced disease at the time of presentation. Distant metastases are not rare. The transfer of novel chemotherapy regimens and preoperative radiation therapy should be considered because pelvic recurrences are the leading cause of progression in SCC.     

Management of multiple echinococcosis in childhood with albendazole and surgery

Background/Purpose

Multiple echinococcosis (ME) is a severe disease in childhood inaccessible to an initial radical surgical treatment. The aim of this study was to evaluate the efficacy of Albendazole in ME and to discuss the role of surgery in this pathology.

Methods

Eleven patients were included in a prospective study between 1996 and 2004. ME was defined by the presence of 10 or more cysts in the same organ. Albendazole was given as 10 mg/kg daily continuously.Treatment outcome was defined as cure, improvement, stabilization or deterioration. Surgery was discussed after 1 year of treatment.

Results

Our patients totalized 296 cysts located essentially on the liver (178 cysts) and the lungs (78 cysts). With exclusive Albendazole therapy, 57.7% pulmonary cysts and 96% peritoneal cysts were considered as cured. This rate was only 31.5% in hepatic localization. After surgical therapy, 67.4% of hepatic cysts were cured. No productive biliary fistula was observed. Two patients were operated laparoscpically. The total treatment duration ranged between 1 and 5 years. Parasitologic examination of operated cysts showed that 30% of them were viable even after 3 years of treatment.

Conclusion

Albendazole has proven a strong efficacy in pulmonary and peritoneal localizations. A complementary surgical treatment is often necessary in hepatic localizations and it is facilitated with previous Albenazole therapy. Combination of ABZ and surgery seems to have encouraging results and must be applied for those patients. The high rate of viable cysts after medical therapy is problematic and must incite to develop new antihelminthic agents.