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Biomedical subjects

J Huguet

Publications and source records attributed to J Huguet.

At least 19 recordsLinked to original sources
PubMed1998

A single step purification, immobilization, and hyperactivation of lipases via interfacial adsorption on strongly hydrophobic supports

A number of bacterial lipases can be immobilized in a rapid and strong fashion on octyl-agarose gels (e.g., lipases from Candida antarctica, Pseudomonas fluorescens, Rhizomucor miehei, Humicola lanuginosa, Mucor javanicus, and Rhizopus niveus). Adsorption rates in absence of ammonium sulfate are higher than in its presence, opposite to the observation for typical hydrophobic adsorption of proteins. At 10 mM phosphate, adsorption of lipases is fairly selective allowing enzyme purification associated with their reversible immobilization. Interestingly, these immobilized lipase molecules show a dramatic hyperactivation. For example, lipases from R. niveus, M. miehei, and H. lanuginosa were 6-, 7-, and 20-fold more active than the corresponding soluble enzymes when catalyzing the hydrolysis of a fully soluble substrate (0.4 mM p-nitrophenyl propionate). Even higher hyperactivations and interesting changes in stereospecificity were also observed for the hydrolysis of larger soluble chiral esters (e.g. (R,S)-2-hydroxy-4-phenylbutanoic ethyl ester). These results suggest that lipases recognize these "well-defined" hydrophobic supports as solid interfaces and they become adsorbed through the external areas of the large hydrophobic active centers of their "open and hyperactivated structure". This selective interfacial adsorption of lipases becomes a very promising immobilization method with general application for most lipases. Through this method, we are able to combine, via a single and easily performed adsorption step, the purification, the strong immobilization, and a dramatic hyperactivation of lipases acting in the absence of additional interfaces, (e.g., in aqueous medium with soluble substrate). Copyright 1998 John Wiley & Sons, Inc.

Journal Article↗
PubMed1998

Human immunodeficiency virus type 1--serodiscordant couples can bear healthy children after undergoing intrauterine insemination.

OBJECTIVE: To use semen from men who were seropositive for human immunodeficiency virus type 1 (HIV-1) to inseminate their partners without infecting them. DESIGN: Prospective study. SETTING: Private practice. PATIENT(S): Sixty-three HIV-1-seropositive men and their HIV-1-seronegative female partners. INTERVENTION(S): The men provided 107 semen samples that were prepared with the use of the Percoll and swim-up techniques. The presence of HIV-1 was determined in the fraction of motile spermatozoa obtained after washing. If HIV-1 was not detected. IUI was performed in stimulated cycles. MAIN OUTCOME MEASURE(S): Human immunodeficiency virus type 1 RNA and DNA were detected with the use of the polymerase chain reaction technique modified for spermatozoa. RESULT(S): One hundred seven semen samples were washed. Human immunodeficiency virus type 1 was not detected in 101 samples (94.4%) and was detected in 6 samples (5.6%). In the latter cases, IUI was not performed. One hundred one IUI procedures were performed in 63 women. Thirty-one pregnancies resulted, for a pregnancy rate of 30.7% per cycle and 49.2% per inseminated woman. Thirty-seven healthy children were born. The results of tests for the detection of HIV-1 and antibodies to HIV-1 in the inseminated women were negative. CONCLUSION(S): On the basis of these results, testing for HIV-1 with the use of the polymerase chain reaction technique on the semen fraction obtained after washing appears to prevent infection in the inseminated woman. This method makes it possible to help HIV-1-seropositive men to have children without infecting their female partners.

Adult↗
PubMed1998

Pregnancy following intracytoplasmic sperm injection from an HIV-1-seropositive man.

The first pregnancy achieved in a seronegative woman following in-vitro fecundation through intracytoplasmic sperm (ICSI) injection from a man with autoimmune deficiency syndrome (AIDS; HIV-1 carrier) is reported. The semen was prepared by PureSperm and swim-up techniques. Some of the motile spermatozoa obtained were used to detect the presence of HIV-1 using the polymerase chain reaction technique. HIV-1 in DNA or RNA form was not detected using this technique. The remaining spermatozoa were frozen. Ovarian stimulation in the woman was performed with long-protocol analogues and gonadotrophins. Thirteen mature oocytes were recovered, into which the thawed spermatozoa were microinjected. Nine embryos were obtained. Four were frozen, four transferred and one discarded. The woman became pregnant. Analyses for HIV-1 in the woman, performed in the first and third months of pregnancy, gave negative results. This case provides further experience with washed semen of sufficient quality for performing artificial insemination in HIV-1-serodiscordant couples (101 inseminations, 31 pregnancies, 28 deliveries, 37 babies, all healthy). In women with obstructed Fallopian tubes, or when the semen is not of sufficient quality for artificial insemination techniques to be performed, ICSI can be carried out using frozen, HIV-1-free semen.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Immunological quantitation of nuclear steroid receptors to optimize the biological classification of breast tumors.

We used immunological methods to determine cytosolic and nuclear steroid receptors to evaluate the advantages of nuclear receptor measurement in the selection of breast cancer patients for treatment. Around 75% of tumors showed coincidence between nuclear and cytosolic receptors (+/+ or -/-) for estrogen receptor (ER) and for progesterone receptor (PgR). Only cytosolic receptors were detected in around 20% of tumors. Distributed in the ER/PgR phenotypes according to the nuclear or cytosolic receptors, 64% of tumors remained in the same subgroup, whereas 16% of tumors were classified as hormone dependent according to cytosolic and independent according to nuclear receptors, which could be considered as 'false-positive' results. 6% of tumors would be classified as negative according to cytosolic receptors but positive according to nuclear receptors and would correspond to 'false-negative' results by conventional methods. Cytosolic receptor results may overrate the hormone dependence and cause some 'misclassifications' of patients. This could partially explain the lack of response to therapy in some cases.

Adult↗
PubMed1997

Associative behaviour of hydrophobically modified carboxymethylpullulan derivatives.

Hydrophobically modified carboxymethylpullulan (HMCMP) samples were obtained by reaction of small amounts of C16 alkylamine on carboxylic groups of the corresponding polyacid. The molar contents of alkyl chains ranged from 1.3 to 6.8% with respect to the anhydroglucose units (AGU) and the degree of substitution (DS) of carboxylic groups varied from 0.76 to 0.84. Solution properties of the sodium salt of HMCMPs were studied mainly by viscometric and size-exclusion chromatography/light-scattering methods. The low-shear viscosity of modified pullulans in 0.1 M NaCl solutions drastically increases with the hydrophobic content and polymer concentrations and the 6.8% modified sample has a quite pseudoplastic behaviour. These data showed that the polymers aggregated intermolecularly and displayed a compact globular structure in dilute solution. Furthermore, addition of NaCl or ethanol induced a decrease in viscosity although the molecular weights remained approximately constant. These results are consistent with a collapse of the polysaccharide aggregates.

Amines↗
PubMed1995

Expression of ecto-adenosine deaminase and CD26 in human T cells triggered by the TCR-CD3 complex. Possible role of adenosine deaminase as costimulatory molecule.

The expression of surface adenosine deaminase (ADA) and CD26 in activated human T cells was studied by flow cytometry. PBLs and CD3+ or CD4+ cells, when subjected to a variety of stimuli (anti-CD3 Abs plus IL-2 or phorbol esters), presented two structurally different cell populations, which differed in size and cellular complexity (populations B1 and B2). In PBLs triggered by an anti-CD3 mAb there was no significant increase of expression of either surface ADA or CD26 in cells of population B1, whose structure is similar to that of nonstimulated cells. In contrast, there was a significant increase in the percentage of expression of ADA and CD26 in the population B2, which corresponds to structurally more complex and larger cells. In the case of activation via TCR-CD3 but in the presence of IL-2 or via phorbol esters, the increase was found in cells from both populations, but B2 cells always showed a higher percentage of expression than B1 cells. The results of increased expression of surface ADA and CD26 were similar in whole T cells or in purer preparations such as CD3+ or CD4+ lymphocytes. Polyclonal Abs against ADA were not able to induce an activation response in T cells even when cross-linked by a secondary Ab. Interestingly, these Abs produced anergy in CD4+ cells subjected to an anti-CD3 stimulus. In contrast, addition of ADA produced an enzyme-independent synergism in the response through the TCR-CD3 complex. In T cells, ADA and CD26 colocalized on the surface of T cells; thus, the effect of exogenous ADA seems to be mediated by CD26 molecules that are not interacting with endogenous ADA (spare CD26 molecules). The presence of spare CD26 molecules on the surface of CD4+ cells was demonstrated by flow cytometry in the presence of exogenous ADA and also by confocal microscopy. The set of results strongly indicates that ADA binding to CD26 produces a costimulatory response in T cell activation events.

Adenosine Deaminase↗
PubMed1995

Surface adenosine deaminase. A novel B-cell marker in chronic lymphocytic leukemia.

Previous studies found that ADA is present on the surface of mononuclear blood cells from healthy patients. Because the expression of this surface antigen depends upon the cell type, the presence of ADA on the plasma membrane of cells from patients with malignant hematologic diseases was studied by flow cytometry. The highest percentage of expression was found in CLL, whereas the lowest was found in T-cell-derived malignancies. The enzyme expression in immortalized cell lines showed a similar pattern, with the highest expression (95% +/- 5%) in the SKW64 B-derived cell line, the lowest (15% +/- 5%) in Jurkat T-lymphoma derived cells, and the intermediate (32% +/- 8%) in K562 cells derived from a chronic myelogenous leukemia. Double labeling ADA/CD5 and ADA/CD19, as well as the correlation of ADA expression with the expression of other surface markers, indicate that surface ADA might be considered a novel marker for CLL.

Adenosine Deaminase↗
PubMed1995

[Open surgery of calculus of the kidney pelvis. Results and complications in the era of extracorporeal lithotripsy].

From 1989 to 1993, 11,535 extracorporeal shock wave lithotripsy (ESWL) sessions were performed in 6,066 patients in our centre. In contrast, simple or extended pyelolithotomy was the treatment of choice in a small group of patients. The object of this paper is to review the results and complications of this type of operation. A total of 57 pyelolithotomies was performed, including 33 extended pyelolithotomies, in 55 patients (26 men and 29 women). The mean age of the series was 50 years (range: 5-77). The operated kidney was the left kidney in 34 cases and the right kidney in 23 cases. The mean interval between the diagnosis and the operation was 31 months. 64% of the patients had history of stone disease (33 ipsilateral and 2 bilateral). Thirteen patients had already been treated by ESWL (10 patients), percutaneous nephrolithotomy (PNL) (2 cases) and one case had previously undergone pyelolithotomy. A urinary tract infection was observed in 26 patients (47%). The incision consisted of lumbotomy in 40 patients (71%), or a subcostal incision in 12 cases. The mean operating time was 161 minutes (range: 90-245) and the mean intraoperative bleeding was 500 ml. Thirteen patients were transfused (23%). The renal artery had to be clamped in 9 cases (for an average of 19 minutes). After a mean follow-up of 20 month, 50.8% of patients were stone-free. 49% of patients presented residual caliceal fragments on the immediate postoperative X-ray. One patient had to be treated by percutaneous nephrolithotomy and ESWL. 12 cases (41% of patients with residual fragments) required subsequent treatment by ESWL and the remaining patients with residual fragments were asymptomatic and did not receive any further treatment. The immediate postoperative complication rate was 17% and 10% of patients developed late postoperative complications. Simple or extended pyelolithotomy remains an effective treatment for stones of the renal pelvis, either on its own or associated with ESWL. However, the possibility of serious complications demands meticulous patient selection.

Adolescent↗
PubMed1993

Surface expression of adenosine deaminase in mitogen-stimulated lymphocytes.

Adenosine deaminase (ADA) expression on the surface of mitogen-stimulated lymphocytes was studied by flow cytometry. The gate for lymphocytes was located by cell size (forward scatter), cytoplasmic complexity (side scatter) and by expression of the markers CD2, CD4, CD8 and CD19. After mitogenic proliferation two populations appeared, one corresponding to non-stimulated cells, and the other consisting of larger cells which showed relatively high expression of adenosine deaminase on their surface. The increase was similar to that observed for CD71 expression, and paralleled the increase in 3H-thymidine incorporation. There was a correlation between ADA and CD71 expression (r = 0.92 for phytohaemagglutinin (PHA) and 0.97 for pokeweed mitogen (PWM)). These results suggest a role for ecto-adenosine deaminase in lymphocyte proliferation and/or triggering.

5'-Nucleotidase↗
PubMed1993

Diagnostic accuracy evaluation using ROC curve analysis.

The diagnostic accuracy of a biochemical quantity is inversely related to the overlapping zone between the values of the population suffering from a disease and the population which does not. The ROC curves are an indirect measure of the overlapping zone between both populations. Specimens (plasma and urine) taken from 928 patients with symptoms of acute abdominal pain were used and the catalytic concentration of alpha-amylase, pancreatic alpha-amylase and triacylglycerol lipase (determined by two methods) were measured. Definitive diagnosis was obtained by following the directives of expert groups on the evaluation of diagnostic tests. Diagnostic accuracy was characterized by calculating the diagnostic sensitivity and specificity, by representing the ROC curves and by quantifying the areas under the ROC curves. The catalytic concentration of pancreatic alpha-amylase in plasma was the quantity with a greater area under the ROC curve (A = 0.9740) and then the one which had greatest diagnostic accuracy. If we considered the upper limit of the reference interval to be the cut-off value, the catalytic concentration of pancreatic alpha-amylase in plasma had a diagnostic sensitivity and specificity values of 0.96 and 0.88 respectively for the acute pancreatitis.

Acute Disease↗
PubMed1991

[Clinical and methodological considerations in the quantification of estrogen and progesterone receptors using monoclonal antibodies in breast cancer].

BACKGROUND: The aim of the present study was to provide information on several clinical and methodological aspects of the measurement of the contents of estrogen (ER) and progesterone (PgR) receptors in breast tumoral tissue using monoclonal antibodies. METHODS: The contents of ER and PgR were measured in 94 tumor specimens of breast carcinoma from pre- and postmenopausal women by the classical method of dextran-charcoal (DC) and an enzyme immunoassay (EIA) with monoclonal antibodies against receptors. RESULTS: The most remarkable results were: a significant difference for ER depending on the menopausal status, an increasing concentration of ER and PgR with age, a greater frequency of the phenotypes ER positive (+)-PgR (+) and ER negative (-) -PgR (-), a lower frequency of positive than negative receptors in the tumors measuring more than 10 cm, a greater presence of tumors with negative than with positive receptors in those located in the lower half of the breast, and the high diagnostic sensitivity of ER and PgR measurement with this method. CONCLUSIONS: There was a remarkable difference in contents of total ER in breast tumors from pre- and post menopausal women. This finding confirms that these tumors may have a different hormone modulation and a different capacity for protein induction depending on the menopausal status.

Adult↗
PubMed1991

Biological variation in the catalytic concentration of pancreatic alpha-amylase and triacylglycerol lipase in serum.

The within-subject and between-subject biological variation in the catalytic concentration of pancreatic alpha-amylase and triacylglycerol lipase in serum have been studied. The study was performed during a 13-month period in a group of 42 supposedly healthy individuals, 19 women and 23 men. All measurements were carried out using a Hitachi 717 analyser at 37 degrees C. The medians of the within-subject biological coefficients of variation obtained for the catalytic concentration of pancreatic alpha-amylase and triacylglycerol lipase in serum were 8.1 and 38.9%, respectively. The index of individuality for the catalytic concentration of pancreatic alpha-amylase and triacylglycerol lipase in serum was 0.27 and 1.20, respectively. Consequently, it is not advisable to use population-based reference intervals for the catalytic concentration of pancreatic alpha-amylase and triacylglycerol lipase in serum.

Adult↗
PubMed1991

A fluorometric enzyme immunoassay for follitropin and lutropin.

The analytical performance of the Stratus (Baxter) automatic analyser for human lutropin and follitropin determination was evaluated and compared with that of an immunoradiometric assay. Within-run and between-run imprecision were lower than those of the immunoradiometric assay. No significant differences were obtained in the parallelism study. A good and significant correlation was obtained between both methods. Results obtained for both methods were not interchangeable. The Stratus automatic analyser can replace radioisotopic methods, thus eliminating radioactive hazards.

Blood Chemical Analysis↗
PubMed1991

[Functional significance of the serum/saliva testosterone ratio in various diseases].

Serum/salivary testosterone ratio (ST/SlvT) expresses the relationship in absolute values between bound and unbound testosterone. This ST/SlvT ratio in supposedly healthy men (n = 25) and women (n = 72) and in patients with several disorders, prostatic carcinoma (n = 19), varicocele (n = 9) and hirsute women (n = 16), has been studied. Both serum and salivary testosterone were measured by an RIA method. ST/SlvT ratio values found in healthy men (78.4 +/- 30.9) did not differ significantly from values found in the varicocele group (111.1 +/- 49.3), but a significant difference (p less than 0.001) from those found in men with prostatic carcinoma (12.3 +/- 7.2) was observed. When the ST/SlvT ratio values obtained in healthy women (18.1 +/- 7.3) were compared with those obtained in hirsute women (1.56 +/- 5.7) no significant differences were observed. The results obtained may indicate the dissociation among the hormone transport, testosterone metabolic clearance and hormone secretion by the salivary glands.

Adult↗