[Consensus documents and clinical guidelines on resistance to antiretroviral agents].
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Anti-HIV Agents↗
Biomedical subjects
L Guerra
Publications and source records attributed to L Guerra.
[GESIDA Consensus document on the use of resistance studies in clinical practice].
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Anti-HIV Agents↗
Activation of A(3) adenosine receptor induces calcium entry and chloride secretion in A(6) cells.
We have previously demonstrated that in A(6) renal epithelial cells, a commonly used model of the mammalian distal section of the nephron, adenosine A(1) and A(2A) receptor activation modulates sodium and chloride transport and intracellular pH (Casavola et al., 1997). Here we show that apical addition of the A(3) receptor-selective agonist, 2-chloro-N(6)-(3-iodobenzyl)-adenosine-5'-methyluronamide (Cl-IB-MECA) stimulated a chloride secretion that was mediated by calcium- and cAMP-regulated channels. Moreover, in single cell measurements using the fluorescent dye Fura 2-AM, Cl-IB-MECA caused an increase in Ca(2+) influx. The agonist-induced rise in [Ca(2+)](i) was significantly inhibited by the selective adenosine A(3) receptor antagonists, 2,3-diethyl-4, 5-dipropyl-6-phenylpyridine-3-thiocarboxylate-5-carboxylate (MRS 1523) and 3-ethyl 5-benzyl 2-methyl-6-phenyl-4-phenylethynyl-1, 4-(+/-)-dihydropyridine-3,5-dicarboxylate (MRS 1191) but not by antagonists of either A(1) or A(2) receptors supporting the hypothesis that Cl-IB-MECA increases [Ca(2+)](i) by interacting exclusively with A(3) receptors. Cl-IB-MECA-elicited Ca(2+) entry was not significantly inhibited by pertussis toxin pretreatment while being stimulated by cholera toxin preincubation or by raising cellular cAMP levels with forskolin or rolipram. Preincubation with the protein kinase A inhibitor, H89, blunted the Cl-IB-MECA-elicited [Ca(2+)](i) response. Moreover, Cl-IB-MECA elicited an increase in cAMP production that was inhibited only by an A(3) receptor antagonist. Altogether, these data suggest that in A(6) cells a G(s)/protein kinase A pathway is involved in the A(3) receptor-dependent increase in calcium entry.
Adenosine↗
Toward epidermal stem cell-mediated ex vivo gene therapy of junctional epidermolysis bullosa.
Junctional epidermolysis bullosa (JEB) is a group of severe, inherited skin diseases caused by mutations in the genes encoding laminin 5 or other components of the hemidesmosome. Since human epidermis is a self-renewing tissue, gene therapy of JEB requires the stable integration of the transgene into the genome of the epidermal stem cell. Human epidermal stem cells can indeed be cultivated and stably transduced with replication-defective retroviral vectors, allowing full phenotypic correction of the adhesion properties of JEB keratinocytes. Epidermal stem cells generate cohesive sheets of stratified epithelium suitable for the permanent coverage of massive skin defects, and genetically modified epidermal sheets maintain long-term expression of the transgene after transplantation on immunodeficient animals. Moreover, we have developed a clinical procedure that allows transplantation of cultured epidermal sheets on large body areas under local anesthesia and without cicatricial outcomes. Thus, (1) the possibility of cultivating lining epithelia, (2) the availability of noninvasive surgical procedures that allow the grafting of large skin areas, and (3) the demonstration of sustained transgene expression in vitro and in vivo by epidermal stem cells, prompt us to propose the implementation of a phase I/II clinical trial aimed at the ex vivo gene therapy of selected JEB patients. The aim of the trial is to validate the ex vivo procedure in a clinical setting, to prove its overall safety, and to analyze critical issues such as long-term survival of the genetically modified implant, immune response against the transgene product, and persistence of transgene expression at therapeutic levels.
Cells, Cultured↗
Phosphoinositide 3-kinase is involved in the tumor-specific activation of human breast cancer cell Na(+)/H(+) exchange, motility, and invasion induced by serum deprivation.
Whereas the tumor acidic extracellular pH plays a crucial role in the invasive process, the mechanism(s) behind this acidification, especially in low nutrient conditions, are unclear. The regulation of the Na(+)/H(+) exchanger (NHE) and invasion by serum deprivation were studied in a series of breast epithelial cell lines representing progression from non-tumor to highly metastatic cells. Whereas serum deprivation reduced lactate production in all three cells lines, it inhibited NHE activity in the non-tumor cells and stimulated it in the tumor cells with a larger stimulation in the metastatic cells. The stimulation of NHE in the tumor cell lines was the result of an increased affinity of the internal H(+) regulatory site of the NHE without changes in sodium kinetics or expression. Serum deprivation conferred increased cell motility and invasive ability that were abrogated by specific inhibition of the NHE. Inhibition of phosphoinositide 3-kinase by overexpression of a dominant-negative mutant or wortmannin incubation inhibited NHE activity and invasion in serum replete conditions while potentiating the serum deprivation-dependent activation of the NHE and invasion. These results indicate that the up-regulation of the NHE by a phosphoinositide 3-kinase-dependent mechanism plays an essential role in increased tumor cell invasion induced by serum deprivation.
Adenosine Triphosphate↗
Treatment of "stable" vitiligo by Timedsurgery and transplantation of cultured epidermal autografts.
OBJECTIVE: To optimize melanocyte/keratinocyte co-cultivation and to evaluate the effectiveness of autologous cultured epidermal grafts in the surgical treatment of stable vitiligo. DESIGN: After optimization of melanocyte/keratinocyte cultures, achromic lesions were disepithelialized by means of programmed diathermosurgery (Timedsurgery) and covered with autologous epidermal grafts prepared from secondary cultures. Melanocyte content was evaluated by dopa reaction. The percentage of repigmentation was calculated using a semiautomatic image analysis system. SETTING: A biosafety level 3 cell culture facility and a dermatological department in a hospital. PATIENTS: Thirty-two patients carrying different types of vitiligo were admitted to the study and treated with autologous cultured epidermal grafts. Inclusion criteria were (1) failure of at least 2 standard medical approaches; (2) no therapy for at least 12 months; (3) absence of progression of old lesions, absence of appearance of new lesions, and absence of Koebner phenomenon within the past 18 months; and (4) absence of autoimmune disorders. RESULTS: One hundred five achromic lesions (a total of 6078.2 cm(2)) were treated. The average percentage of repigmentation, evaluated after 12 to 36 months of follow-up, was 77%. Independent of the type of vitiligo, average percentages of repigmentation of extremities and periorificial sites were 8% (31.8 cm(2) repigmented/420.5 cm(2) transplanted) and 35% (17.6 cm(2) repigmented/50.0 cm(2) transplanted), respectively. Percentages of repigmentation of all other body sites ranged from 88% to 96% (4329.7 cm(2) repigmented/4675.2 cm(2) transplanted). Color matching was good and scar formation was not observed. CONCLUSION: Cultured epidermal grafts can be considered a real therapeutic surgical alternative for "stable" but not lip-tip vitiligo.
Adolescent↗
Cytokine analysis before and after mycophenolate mofetil treatment in renal transplantation.
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Antigens, CD↗
Thermodynamically distinct high and low affinity states of the A(1) adenosine receptor induced by G protein coupling and guanine nucleotide ligation states of G proteins.
The influence of the receptor-G protein coupling state and the guanine nucleotide ligation state of the G protein on the binding mechanism of A(1) adenosine receptor ligands has been investigated in [(3)H]-1,3-dipropyl-8-cyclopentylxanthine ([(3)H]-DPCPX) binding studies in rat brain membranes. Thermodynamic parameters of binding of A(1) adenosine receptor ligands of different intrinsic activities were determined in the absence or presence of GDP and compared to the binding mechanism after receptor-G protein uncoupling. In agreement with previous studies, it was found that xanthine and non-xanthine antagonists showed an enthalpy- or enthalpy- and entropy-driven binding mechanism under all conditions. In contrast to antagonists, the binding mechanism of agonists was strongly affected by the G protein coupling state or the absence or presence of guanine nucleotides. Binding of full and partial agonists to the high-affinity state of the A(1) receptor was entropy-driven in the absence of GDP, and a good correlation between intrinsic activities and the contribution of entropy was observed. In the absence of GDP, binding of full and partial agonists and antagonists to the high affinity state of the receptor was thermodynamically discriminated. In contrast, no such discrimination was found in the presence of GDP. The binding mechanism of agonists to the low-affinity state of the receptor was identical to that of antagonists only after uncoupling of the receptor from G proteins by pretreatment with N-ethylmaleimide or guanosine-5'-(gamma-thio)-triphosphate (GTPgammaS). These results indicate the existence of two thermodynamically distinct high- and low-affinity states of the A(1) adenosine receptor.
Animals↗
Assessment of antibiotic prescription in acute respiratory infections in adults. The Spanish Study Group on Antibiotic Treatments.
OBJECTIVE: This study aims to ascertain the variability in the use of antibiotics for the treatment of acute respiratoryinfections in several hospital emergency services in Spain, as well as the appropriateness of antibiotics prescription through evaluation by a panel of experts using available scientific evidence. METHOD: A cross-sectional study was carried out in the emergency services of 10 hospitals in different Spanish regions. We chose patients diagnosed as having acute respiratory infection, aged over 14 years. Among the collected variables were: type of respiratory infection, antibiotic prescription, comorbidity, qualification of the prescribing doctor and hospital admission. The consensus conference held by a panel of experts established first choice treatment and the alternative and inappropriate use for each respiratory infection, based on the available scientific evidence. All the observed prescriptions in our study were classified according to this pattern. RESULTS: A sample of 2899 acute respiratory infections was studied (5.5% of all emergencies). Antibacterial agent treatment was prescribed in 82.6% of these, varying according to the infection between 98.5% of pneumonias and 49% of croup-influenza-common cold. The most commonly used antibiotics were amoxicillin-clavulanate and cefuroxime. The global percentage of inappropriate prescription was 40.5% (95% CI; 35.4-45.5). The prescriptions were inappropriate in 16.9% of cases of pharyngotonsillitis, 17.8% of chronic bronchitis, 26.9% of acute bronchitis, 29.3% of pneumonias, 30.8% of otitis and sinusitis and in 70.8% of croup, flu, common cold and non-specified infections. Significant variability among participating centres was observed, both in choice of antibiotics and in their degree of appropriateness. CONCLUSIONS: There is excessive use of antimicrobial drugs in acute respiratory infections, and the majority are used for viral infections. There is indiscriminate use of broad spectrum antibiotics, which are valid in some infections but clearly inappropriate in others. Similarly, there are important differences in the choice of antibiotics and their degree of appropriateness among hospitals.
Acute Disease↗
Prothrombin gene mutation uncommon in pulmonary embolism.
BACKGROUND: Venous thrombosis followed by pulmonary embolism is one of the most common causes of sudden death among middle-aged adults. Several inherited polymorphisms are associated with heightened risk of venous thrombosis, including mutation at position 20210 of the prothrombin gene and mutation at codon 506 of the factor V gene. METHODS: We studied mutation prevalence in 67 individuals who died of pulmonary embolism and were autopsied in a medical examiner's facility over a 5-year period. Mutations were identified by polymerase chain reaction followed by allele-specific endonuclease digestion. RESULTS: Traditional risk factors for pulmonary embolism (eg, immobility, oral contraceptive use, cancer) were identified in 75%. Heterozygous mutation of the prothrombin gene was found in 3/67 (4%), and heterozygous mutation of the factor V gene was identified in 3/66 (4%). No homozygotes or compound heterozygotes were identified. The prevalence of mutation was not significantly different from that of the general population. CONCLUSIONS: Individuals who die suddenly from pulmonary embolism are not often affected by prothrombin or factor V gene mutations. Therefore, medical examiners need not routinely test for these mutations in individuals who die of pulmonary embolism.
Adolescent↗
[Recommendations of GESIDA (Grupo de Estudio de SIDA)/National Plan on AIDS with respect to the anti-retroviral treatment in adult patients infected with the human immunodeficiency virus in the year 2000 (II)].
OBJECTIVE: To update the recommendations for antiretroviral therapy (ART) in adult HIV-infected persons according to the new scientific advances and the existence of new antiretroviral drugs in the last two years. METHODS: The ART recommendations have been condensed by a panel of experts from the Spanish AIDS Study Group (Grupo de Estudio de Sida-GESIDA) of the Spanish Infectious Diseases and Clinical Microbiology Society (SEIMC) and from the Clinical Advisory Panel (CAP) of the Secretariat of the Spanish National Plan on AIDS (SPNS) of the Ministry of Health. Three levels of evidence have been established depending if the data came from randomized and controlled studies, from cohort or case-control studies or from descriptive studies and expert opinions, for that purpose we have reviewed the advanced in HIV pathophysiology and results of efficacy (clinical, virologic and immunologic) and security (toxicity) from clinical trials involving ART lasting at least 12 months, from cohort studies and pharmacokinetic and security data of antoiretrovírico drugs, presented in international conferences or published in biomedical journals in the last two years. In each situation we have established either to recommend or to consider or not recommend ART. RESULTS: Nowadays, ART consistent of at least three drugs constitutes the election therapy for chronic HIV infection, since it delays clinical progression, increases significantly the survival and diminishes hospital admissions and associated costs. The decision to start ART must be based upon three elements: presence or absence of symptoms, plasma vírica load and CD4+ cells counts. Thus, in asymptomatic cases with a high CD4+ cells count (> 500/microliter) and low vírica load (< 10,000 copies/ml by branched DNA bDNA or < 20,000 copies/ml by reverse-transcription polymerase chain reaction [RT-PCR] or nucleic acid sequence based amplification [NASBA]) we recommend to delay ART. In symptomatic patients we recommend to start it, and in asymptomatic patients, we could recommend or consider ART initiation depending on the risk of progression, established by the vírica load and the CD4+ cells count. In any case, if therapy is started, the objective must be to reach an indetectable vírica load (< 50 copies/ml). The adherence to ART plays a key role for its initial moment and for the duration of the antiviral response. ART can achieve a restoration of cellular immunity inb the advanced patients. There are few therapeutic options in failing patients due to cross-resistance. Resistance studies can be useful in this setting. The toxicity (lypodistrophy) is a new and limiting factor of ART which requires to look for new therapeutic options. ART criteria for acute infection, pregnancy, post-exposure prophylaxis and when to use resistance testing are discussed. CONCLUSIONS: In this moment, there is a more conservative attitude towards starting ART than in previous recommendations in which a virus eradication was considered. On the other hand, the high number of disposable drugs, the more sensitive monitorization methods (plasma vírica load) and the possibility of performing resistance studies make therapeutic strategies more dynamic and individualized for each patient and situation. In any case, it is mandatory to ensure a perfect adherence to ART from the patients.
Acquired Immunodeficiency Syndrome↗
The control of epidermal stem cells (holoclones) in the treatment of massive full-thickness burns with autologous keratinocytes cultured on fibrin.
BACKGROUND: Cell therapy is an emerging therapeutic strategy aimed at replacing or repairing severely damaged tissues with cultured cells. Epidermal regeneration obtained with autologous cultured keratinocytes (cultured autografts) can be life-saving for patients suffering from massive full-thickness burns. However, the widespread use of cultured autografts has been hampered by poor clinical results that have been consistently reported by different burn units, even when cells were applied on properly prepared wound beds. This might arise from the depletion of epidermal stem cells (holoclones) in culture. Depletion of holoclones can occur because of (i) incorrect culture conditions, (ii) environmental damage of the exposed basal layer of cultured grafts, or (iii) use of new substrates or culture technologies not pretested for holoclone preservation. The aim of this study was to show that, if new keratinocyte culture technologies and/or "delivery systems" are proposed, a careful evaluation of epidermal stem cell preservation is essential for the clinical performance of this life-saving technology. METHODS: Fibrin was chosen as a potential substrate for keratinocyte cultivation. Stem cells were monitored by clonal analysis using the culture system originally described by Rheinwald and Green as a reference. Massive full-thickness burns were treated with the composite allodermis/cultured autograft technique. RESULTS: We show that: (i) the relative percentage of holoclones, meroclones, and paraclones is maintained when keratinocytes are cultivated on fibrin, proving that fibrin does not induce clonal conversion and consequent loss of epidermal stem cells; (ii) the clonogenic ability, growth rate, and long-term proliferative potential are not affected by the new culture system; (iii) when fibrin-cultured autografts bearing stem cells are applied on massive full-thickness burns, the "take" of keratinocytes is high, reproducible, and permanent; and (iv) fibrin allows a significant reduction of the cost of cultured autografts and eliminates problems related to their handling and transportation. CONCLUSION: Our data demonstrate that: (i) cultured autografts bearing stem cells can indeed rapidly and permanently cover a large body surface; and (ii) fibrin is a suitable substrate for keratinocyte cultivation and transplantation. These data lend strength to the concept that the success of cell therapy at a clinical level requires cultivation and transplantation of stem cells. We therefore suggest that the proposal of a culture system aimed at the replacement of any severely damaged self-renewing tissue should be preceded by a careful evaluation of its stem cell population.
Adolescent↗
[Evaluation of the quality of life in patients with HIV and AIDS infection].
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Acquired Immunodeficiency Syndrome↗
Alpha subunit of glycoprotein hormones in the sera of acromegalic patients and its mRNA in the tumors.
Within a population of 16 pituitary adenomas we found high levels of glycoprotein alpha subunits in the sera of patients with somatotrophic tumors. This finding was correlated with the presence of mRNA alpha subunit in these tumors indicating the adenomas themselves as the origin of the circulating alpha-subunit. Synthesis of these two hormones, which are chemically very different, by the same tumor cells indicates a high degree of differentiation of these cells. We are unable at this time to conclusively correlate differentiation of these tumors aggressively.
Acromegaly↗
[Effects of the chronic malnutrition on the transferrin receptors in patients with nutritional anemia].
It has been postulated that the provision of iron to the erythroid cells is under the control of a fraction of free intracellular iron. Variations in the size of this pool would send messages to either seric or cellular iron receptors aimed to keep a balance between the offer and the demand by the cells. The possibility that cell membrane receptors could be affected by chronic malnutrition was explored in these studies by the changes of iron uptake capacity of circulating mouse reticulocytes cause by the incubation of the cells with serum of either normal or anemic patients plasma donors. The results show that the incubation with nutritional anemic serum caused a significant drop in iron uptake capacity of the cells. The mechanism of the difference is discussed.
Adolescent↗
[Multicenter study of the variability and adequacy of antimicrobial therapy for community-acquired pneumonia in adults].
We performed a study to evaluate the variability and adequacy of prescribing antibiotics in community-acquired pneumonia (CAP) in 10 Spanish hospitals. We studied 452 patients with CAP. Initial empirical administration of antibiotics was prescribed in 90.7% of the cases, 82.5% as monotherapy. Macrolides and third and second generation cephalosporins were the most widely used groups of antibiotics. Penicillin and amoxicillin were only prescribed in 1. 7% of the patients. A significant variability between hospitals was observed. Reference patterns for the use of antibiotics in CAP were devised by a panel of experts. According to the recommendations of this panel, 29% of the total prescriptions were not adequate, with this percentage reaching 65% in outpatients older than 65 years or with comorbidity. This was mainly due to the fact that monotherapy with erythromycin, which was considered inadequate, was the most widely prescribed treatment.
Adult↗
[Multicenter study of the variability and adequacy of antimicrobial therapy for community-acquired pneumonia in adults]
We performed a study to evaluate the variability and adequacy of prescribing antibiotics in community-acquired pneumonia (CAP) in 10 Spanish hospitals. We studied 452 patients with CAP. Initial empirical administration of antibiotics was prescribed in 90.7% of the cases, 82.5% as monotherapy. Macrolides and third and second generation cephalosporins were the most widely used groups of antibiotics. Penicillin and amoxicillin were only prescribed in 1.7% of the patients. A significant variability between hospitals was observed. Reference patterns for the use of antibiotics in CAP were devised by a panel of experts. According to the recommendations of this panel, 29% of the total prescriptions were not adequate, with this percentage reaching 65% in outpatients older than 65 years or with comorbidity. This was mainly due to the fact that monotherapy with erythromycin, which was considered inadequate, was the most widely prescribed treatment.
Journal Article↗
Cultivation of human keratinocyte stem cells: current and future clinical applications.
Cultured human keratinocytes have a wide spectrum of clinical applications. Clinical results reported by several investigators are, however, contradictory. In this review, the authors discuss the biological and surgical issues which play a key role in the clinical outcome of cultured epidermal autografts used for the treatment of massive full-thickness burns. The importance of cultivation of epidermal stem cells and of their transplantation onto a wound bed prepared with donor dermis is emphasised. The paper also reviews recent data showing that: (i) cultured epidermal autografts bearing melanocytes can be used for the treatment of stable vitiligo; (ii) keratinocytes isolated from other lining epithelia, such as oral, urethral and corneal epithelia, can be cultivated and grafted onto patients suffering from disabling epithelial defects; (iii) keratinocyte stem cells can be stably transduced with retroviral vectors and are therefore attractive targets for the gene therapy of genodermatoses.
Burns↗