Regiospecific Oxidation of Chlorobenzene to 4-Chlororesorcinol, Chlorohydroquinone, 3-Chlorocatechol and 4-Chlorocatechol by Engineered Toluene o-Xylene Monooxygenases.

Toluene o-xylene monooxygenase (ToMO) was found to oxidize chlorobenzene to form 2-chlorophenol (2-CP, 4%), 3-CP (12%), and 4-CP (84%) with a total product formation rate of 1.2 ± 0.17 nmol/min/mg protein. It was also discovered that ToMO forms 4-chlorocatechol (4-CC) from 3-CP and 4-CP with initial rates of 0.54 ± 0.10 and 0.40 ± 0.04 nmol/min/mg protein, respectively, and chlorohydroquinone (CHQ, 13%), 4-chlororesorcinol (4-CR, 3%), and 3-CC (84%) from 2-CP with an initial product formation rate of 1.1 ± 0.32 nmol/min/mg protein. To increase the oxidation rate and alter the oxidation regiospecificity of chloroaromatics, as well as to study the roles of active site residues L192 and A107 of the alpha hydroxylase fragment of ToMO (TouA), we used the saturation mutagenesis approach of protein engineering. Thirteen TouA variants were isolated, among which some of the best substitutions uncovered here have never been studied before. Specifically, TouA variant L192V was identified which had 1.8-, 1.4-, 2.4-, and 4.8-fold faster hydroxylation activity toward chlorobenzene, 2-CP, 3-CP, and 4-CP, respectively, compared to the native ToMO. The L192V variant also had the regiospecificity of chlorobenzene changed from 4% to 13% 2-CP and produced the novel product 3-CC (4%) from 3-CP. Most of the isolated variants were identified to change the regiospecificity of oxidation. For example, compared to the native ToMO, variants A107T, A107N, and A107M produced 6.3-, 7.0-, and 7.3-fold more 4-CR from 2-CP, respectively, and variants A107G and A107G/L192V produced 3-CC (33 and 39%, respectively) from 3-CP whereas native ToMO did not. IMPORTANCE Chlorobenzene is a commonly used toxic solvent and listed as a priority environmental pollutant by the US Environmental Protection Agency. Here, we report that Escherichia coli TG1 cells expressing toluene o-xylene monooxygenase (ToMO) can successfully oxidize chlorobenzene to form dihydroxy chloroaromatics, which are valuable industrial compounds. ToMO performs this at room temperature in water using only molecular oxygen and a cofactor supplied by the cells. Using protein engineering techniques, we also isolated ToMO variants with enhanced oxidation activity as well as fine-tuned regiospecificities which make direct microbial oxygenations even more attractive. The significance of this work lies in the ability to degrade environmental pollutants while at the same time producing valuable chemicals using environmentally benign biological methods rather than expensive, complex chemical processes.

Estrogen receptor alpha mediates mandibular condylar cartilage growth in male mice.

OBJECTIVES: In the appendicular skeleton, estrogen via ERα signalling has been shown to mediate endochondral growth plate fusion in both males and females. However, the role of ERα in mediating growth of the mandibular condylar cartilage is unknown. Thus, this study focuses on the characterization of the mandibular condylar cartilage phenotype in young and adult male ERαKO mice. SETTING: Columbia University Medical Center. MATERIAL AND METHODS: WT and ERαKO C57BL/6 male mice were sacrificed at 49 days or 9 months for phenotypic analysis. Changes to MCC thickness, cell number and cell density were measured using histomorphometric methods. Cartilage-specific gene expression and OARSI scores were investigated for 49-day and 9-month-old male ERαKO and WT mice. RESULTS: In young mice, a significant increase in the number of mandibular condylar cartilage cells and a significant decrease in the expression of Col10, Runx2 and DMP1 were observed in the male ERαKO mice compared to WT. In 9-month-old mice, we found a similar increase in the number of cells but no change in osteoarthritic histological scoring in ERαKO mice compared to WT mice. CONCLUSION: In summary, estrogen plays a role in mediating mandibular condylar maturation in young male mice. However, according to this study, it does not play a role in mediating long-term growth or age-related mandibular condylar cartilage degeneration in males.

Priorities for cancer control in Spain.

Spanish cancer strategy is being developed in the context of a decentralised health care system. The advantages and challenges posed by this system are reviewed, particularly vis-à-vis the need to build consensus among regional health services as regards priorities in cancer control. Analysis of the impact of cancer in Spain enables smoking prevention and obesity reduction to be highlighted, especially among adolescents, as targets for primary prevention. Two-yearly colorectal cancer screening using the faecal occult blood test and targeting persons aged 50-69 years is a new goal that should be pursued countrywide, as should population-based breast cancer screening. Insofar as health care is concerned, the focus is on promoting and consolidating multidisciplinary cancer care based on evidence-based clinical guidelines, and on ensuring that cancer patients enjoy prompt access to diagnosis and therapy. Psychosocial support, with experience gained in several health services, should be expanded. Palliative care must be consolidated as a necessary therapy for patients who require it. Finally, the important boost to cancer research witnessed in recent years should be sustained. The approach adopted is based on building a co-operative strategy among all of Spain's autonomous regions (Comunidades Autónomas), which must then apply the interventions in their respective health services. A shared understanding of the main priorities for cancer control, through a review of the evidence and a consensus embracing all stakeholders, including scientific societies and patients' associations, is an essential step in the implementation of cancer strategy in Spain.

[Herpetic recurrent upper limb lymphangitis]. / Lymphangite récurrente du membre supérieur d'origine herpétique.

Upper limb lymphangitis often complicates varied wounds on the hand or forearm and improvement is obtained in a few days with adapted antibiotic therapy. A 28-year-old woman presented since few years episodes of lymphangitis of the arm associated with vesicles on an erythematous base, on the palmar face of the first phalanx of the index finger, spontaneous relief within 10 days, without antibiotic therapy. Herpetic origin was confirmed on viral culture. No primary infection neither recurrence was noted. Because of the recurrences, a prophylactic treatment with valaciclovir was instituted. There was no reported recurrence at two years follow-up. Upper limb lymphangitis rarely complicates herpetic whitlow in immunocompetent patient. Clinicians should be aware of viral lymphangitis, which is often overlooked and associated with diagnostic errors and treatment delay.

Head-and-neck swelling: an under-recognized feature of giant cell arteritis. A report of 37 patients.

OBJECTIVE: To describe the frequency of occurrence and characteristics of head-and-neck swelling (HNS) in temporal (giant cell) arteritis (TA). METHODS: We analyzed the charts of patients with HNS retrieved from a single department series of 260 consecutive patients with TA and reviewed the published French-English literature. Patients with a swelling limited to the temporal fossa were excluded. RESULTS: A history of HNS was elicited in 17 patients in our series (i.e. 6.5%) and in 20 previously published patients. The swelling was an inaugural feature in most cases and was often transient. Ear-nose-and-throat (ENT )symptoms were observed in 80% of the cases, including jaw claudication or pain upon opening mouth in 22, causing trismus in 10. Two patients had permanent visual impairment and 1 had sudden hearing loss. The temporal artery biopsy yielded giant cell arteritis (GCA) in all the patients but 2. The HNS was often painful and mainly involved mainly the orbital region and face, particularly the lower part of the cheeks and maxillae, less often the neck and, rarely, the forehead and tongue. Concurrent localized limb swellings were also observed in 3 patients. The HNS disappeared in all the patients, either spontaneously or under steroid treatment, and recurred only in 6 patients. CONCLUSION: HNS is not exceptional in untreated TA and is strongly associated with ENT symptoms and a positive TAB, but not with visual loss or stroke. Such characteristics imply in these cases a prominent, widespread involvement of the external carotid artery system by giant cell arteritis.

Aortic angiosarcoma clinically mimicking polyarteritis nodosa.

We report two patients in whom angiosarcoma of the aorta (ASA) with distal emboli and skin metastases was initially clinically misdiagnosed as polyarteritis nodosa (PAN). Both presented with features highly suggestive of systemic necrotizing vasculitis, including constitutional symptoms, orchitis (in one), leg pain with sensory neuropathy, livedo, focal areas of skin necrosis, painful cutaneous nodules demonstrating vasculitis or inconspicuous changes on biopsy, raised acute phase reactants, and transient improvement with systemic corticosteroids. Repeated skin biopsies revealed an atypical endovascular cell proliferation with strong reactivity for factor VIII. Magnetic resonance angiography (MRA) demonstrated an intra-luminal aortic tumour at the infra-renal level. In retrospect, the (PAN)-like features appeared to involve exclusively the lower part of the body in these patients.

Regresión espontánea tras nefrectomía de carcinoma renal metastásico. Aportación de un caso con confirmación histológica y revisión de la literatura / Spontaneous regression of metastatic renal carcinoma after nephrectomy. Presentation of a case with histologic confirmation

El carcinoma renal suele manifestarse clínicamentede forma tardía, por lo que aproximadamenteun tercio de los pacientes presentan metástasis adistancia en el momento del diagnóstico. En estoscasos, la supervivencia disminuye drásticamente, situándoseen una media de 12 meses. Se han descritoalgunos casos de regresión espontánea (RE) en estetumor, que varían desde regresiones completas yduraderas hasta parciales y temporales. La RE sepresenta entre el 0,4 y el 4% de los pacientes concarcinoma renal mestastásico (CRM). A pesar deestar descritos muchos factores relacionados con esteraro y favorable suceso, no se conoce su verdaderoorigen y no es, por tanto, un evento predecible.Presentamos un caso de RE tras nefrectomía en unpaciente con carcinoma renal, que presentaba metástasispulmonares confirmadas histológicamente yrecidiva local

The clinical manifestations of renal cell carcinomaare often late, so that approximately one third ofthe patients have metastatic disease at the time ofdiagnosis. In this case, the survival rate diminishesdramatically, with an average survival time of 12months. However, some cases of spontaneous regressionhave been reported for this kind of tumor,either complete and permanent or partial and temporary.Spontaneous regression occurs in 0.4-4.0%of the patients with metastatic renal cell carcinoma.Many factors have been proposed as related withthis rare but favorable event, but until now the originremains unknown and no prediction can be made.We present a case of renal cell carcinoma withhistologic confirmation of lung metastases and localrelapse showing a spontaneous regression afternephrectomy

[Adverse drug related events in a postemergency unit: prospective cohort study with 6 months follow up]. / Iatrogénie chez le sujet âgé de plus de 75 ans dans un service de posturgences. Etude prospective de cohorte avec suivi à six mois.

PURPOSE: To analyse iatrogenic events in elderly people and determine the part of unplanned admission in postemergency units directly related to thus iatrogenic events. METHODS: The authors conducted a prospective chart review on treatments and potentials adverse drug-related events of all elderly consecutively hospitalized between January and Marsh 2003 in a postemergency department. A 6 months prospective evaluation after discharge was made for all elderly with adverse drug-related event. RESULTS: One hundred (and) eighty-six elderly (mean age 83+/-5.7 years) were prospectively included. Eighty-one per cent are ambulatory with a self-medication administration in spite of a real disability (activity of daily-living: 4.5+/-1.8). The number of medications consumed ranged from 0 to 15 and averaged 6, with to different source of prescriptions in 34% of the cases. The treatment was recently modified in 41 cases (22%). Adverse drug related events accounted in 55 cases (29%) and hospitalization was directly related to iatrogenic event in 32 cases (17%). Adverse drug related events could be avoided in half cases. There was no death directly related with adverse drug reactions. Follow up after discharge was obtained in 47 cases and pointed out elderly disability: 34 were again hospitalized, 14 admitted in nursing home facilities and 12 died. Treatment was equivalent to our prescription only in 35% of the cases; on the other hand, we found only four elderly with medication directly related to previous adverse event. DISCUSSION: Theses results pointed out once again polymedication observed in frail elderly people leading to extreme difficulty to prescription due to polypathology. Prescription renewal could be related to adverse drug related events and precipitated elderly people in disability leading to institutionalization.

[Disease associations in 250 patients with temporal (giant cell) arteritis]. / Maladie de Horton: associations morbides chez 250 malades.

INTRODUCTION: Miscellaneous disorders have been described in association with temporal (giant cell) arteritis (TA), most often anecdotally, except with arteriosclerosis. METHOD: In a retrospective study, we reported our personal experience of disease associations in a series of 250 patients diagnosed with TA and followed-up in the department between 1976 and 2003. RESULTS: Disease associations were found in 43 patients, i.e. 17% of cases: concurrent malignancy (23 patients: 17 cancers and 6 blood diseases), primary Gougerot-Sjögren's syndrome (6 cases), endocrine disease other than Hashimoto's thyroiditis (7 cases: 3 hyperparathyroidism [HPP], 3 hyperthyroidism, 1 association HPP + hyperthyroidism), polyneuropathy (3 cases), essential thrombocythaemia (2 cases), anti-neutrophilic cytoplasmic (anti-myeloperoxidase) antibodies (2 cases), and miscellaneous associations (1 case of RS3PE syndrome, nephrotic syndrome, myasthenia, sarcoidosis, and macro-creatine kinase type 2). More than one disease associated was present 5 patients. In 77% of the patients, there was a strong temporal association between TA and the alternate illness. No systemic necrotizing vasculitis or rheumatoid arthritis was observed in any patient. CONCLUSION: In our experience, there was a frequent, non-fortuitous, association between TA and malignancy. Auto-immune conditions were rare, but the prevalence of Gougerot-Sjögren's syndrome might have been underestimated. Hyperthyroidism and HPP are not exceptional and must be recognised in order to avoid severe bone loss induced by corticosteroids.

[Carcinosarcoma of the bladder: report of our cases and review of the literature]. / Carcinosarcoma vesical: análisis de nuestra serie y revisión de la literatura.

Carcinosarcomas of the bladder are rare. As a result, the natural history of them and the best methods of treatment remain uncertain. These tumors tend to be rapidly growing, invasive and recur locally. We reviewed our experience with four patients presenting between 1995 and 2002. Epidemiology findings, clinical aspects, histological features, diagnostic methods, treatment and survival are discussed in relation to literature.

Análisis de 18 pacientes diagnosticados de distrofia muscular de tipo Becker / Analysis of 18 cases of Becker muscular dystrophy

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Carcinosarcoma vesical: análisis de nuestra serie y revisión de la literatura / Carcinosarcoma of the Bladder: Report of our cases and review of the literature

El carcinosarcoma vesical es una neoplasia de muy baja frecuencia de aparición. Por ello su historia natural y las formas de tratamiento no están claramente determinadas. Suelen ser tumores rápido crecimiento, invasivos y con tendencia a la recurrencia. Presentamos la revisión de los cuatro casos que hemos tenido ocasión de diagnosticar y tratar entre 1995 y 2002. Analizamos los aspectos epidemiológicos, sintomatología presentada, hallazgos histológicos, métodos diagnósticos, tratamiento y supervivencia, revisando además la literatura al respecto (AU)

Carcinosarcomas of the bladder are rare. As a result, the natural history of them and the best methods of treatment remain uncertain. These tumors tend to be rapidly growing, invasive and recur locally. We reviewed our experience with four patients presenting between 1995 and 2002.Epidemiology findings, clinical aspects, histological features, diagnostic methods, treatment and survival are discussed in relation to literature (AU)

[Dysmetabolic hepatosiderosis, characteristics in 51 patients]. / Hépatosidéroses dysmétaboliques, caractéristiques chez 51 patients.

OBJECTIVE: Dysmetabolic hepatosiderosis is a recently described entity, which reunites iron overloads associated with polymetabolic syndromes, steatosis and steato-hepatitis. The aim of this study was to specify the relationship between iron overload, polymetabolic syndrome, HFE mutations of primary hemochromatosis and steatosis. METHOD: This was a 5-year retrospective study of 51 patients hospitalised and/or seen in consultation, non-alcoholic, presenting with hyperferritinemia associated with a polymetabolic syndrome and/or hepatic steatosis. RESULTS: Patients mean age was of 53 +/- 12 years with a sex ratio M/F of 5.4. Metabolic disorders were found in 97% of the non-steatosis patients (overweight: 40%, perturbed sugar metabolism: 47%, dyslipidemia: 79.5%). Hyperferritinemia was constant but moderate (513.4 +/- 280.3 ng/ml). Transferrin saturation was predominantly normal (0.44 +/- 0.17). The hepatic enzymes were normal or only slightly perturbed. Steatosis was revealed by sonography in 62% of cases. The hepatic iron overload, documented in 6 patients by hepatic needle biopsy, was discreet (concentration/age ratio of 1.64 +/- 0.19). Among the 59% patients screened for the HFE gene, 40% were positive for C282Y and/or H63D. There was no correlation between ferritin levels and iron parameters, polymetabolic syndrome parameters and hepatic enzymes. The HFE mutations had no influence on the iron parameters. CONCLUSION: Dysmetabolic hepatosiderosis must be know by hospital practitioners because of their prevalence in cases of hyperferritinemia and their therapeutic incidence.

[Systemic lupus erythematosus and herpes virus infection: three new observations]. / Lupus érythémateux disséminé et herpes viridae: à propos de trois observations.

INTRODUCTION: Systemic lupus erythematosus is still of unknown origin. Viruses have long been postulated to play a role in its pathogenesis particularly cytomegalovirus and Epstein-Barr virus. EXEGESE: We describe three patients who presented acute onset of systémic lupus erythematosus concurrently with recent viral infection (two with cytomegalovirus and one with Epstein-Barr virus). CONCLUSION: The peculiar clinical events emphasize the difficulty of diagnosis at the onset of the disease and suggest possible role of these viruses in the pathogenesis of SLE.