Cytoreductive treatment in patients with CALR-mutated essential thrombocythaemia: a study comparing indications and efficacy among genotypes from the Spanish Registry of Essential Thrombocythaemia.

The present study assessed the criteria for initiating cytoreduction and response to conventional therapies in 1446 patients with essential thrombocythemia (ET), 267 (17%) of which were CALR-mutated. In low risk patients, time from diagnosis to cytoreduction was shorter in CALR-positive than in the other genotypes (2·8, 3·2, 7·4 and 12·5 years for CALR, MPL, JAK2V617F and TN, respectively, P < 0·0001). A total of 1104 (76%) patients received cytoreductive treatment with hydroxycarbamide (HC) (n = 977), anagrelide (n = 113), or others (n = 14). The estimated cumulative rates of complete haematological response (CR) at 12 months were 40 % and 67% in CALR and JAK2V617F genotypes, respectively. Median time to CR was 192 days for JAK2V617F, 343 for TN, 433 for MPL, and 705 for CALR genotypes (P < 0·0001). Duration of CR was shorter in CALR-mutated ET than in the remaining patients (P = 0·003). In CALR-positive patients, HC and anagrelide had similar efficacy in terms of response rates and duration. CALR-mutated patients developed resistance/intolerance to HC more frequently (5%, 23%, 27% and 15% for JAK2V617F, CALR, MPL and TN, respectively; P < 0·0001). In conclusion, conventional cytoreductive agents are less effective in CALR-mutated ET, highlighting the need for new treatment modalities and redefinition of haematologic targets for patients with this genotype.

Prevalence of multiple sclerosis in Santiago de Compostela (Galicia, Spain).

INTRODUCTION: Multiple sclerosis (MS) is the most common central nervous system disease in young adults, and one of the leading causes of disability in this age group. OBJECTIVES: To establish the prevalence and incidence of MS in Santiago de Compostela (Spain). Material and methods We performed a prospective, case-finding study on > 90,188 inhabitants, from 1 January 1998 to 15 September 2003, selected as day of prevalence. Sources of information were the University Clinical Hospital of Santiago, the hospitals and private clinics of the city, rehabilitators, pediatricians, ophthalmologists, psychiatrists, neurosurgeons, specialists in internal medicine, primary care physicians, association of patients of MS, social services and nongovernmental and religious associations. Informative days were programmed for the susceptible population, and campaigns were carried out in the media. All patients with Poser diagnostic criteria for MS, on the census of Santiago de Compostela as of 15 September 2003, were included in the study. RESULTS: On the day of prevalence, 71 patients with MS were registered on the census of Santiago de Compostela, therefore, the prevalence was 79 cases per 100,000 inhabitants. The incidence of the disease in the study period was 5.3 cases per 100,000 inhabitants and year. CONCLUSIONS: The prevalence and incidence rates of MS found in Santiago de Compostela are the highest reported, to date, in Spain.

Isolation of viral hemorrhagic septicemia virus from Greenland halibut Reinhardtius hippoglossoides caught at the Flemish Cap.

Viral hemorrhagic septicemia virus (VHSV) was isolated from apparently healthy Greenland halibut Reinhardtius hippoglossoides caught in the Flemish Cap, a deep fishing ground in the North Atlantic Ocean in international waters near Newfoundland. The identity of the virus was confirmed by electron microscopy, immunodot, seroneutralization and reverse transcriptasepolymerase chain reaction. In the serology assays, all isolates reacted in the immunodot assay with a polyclonal antiserum against the European VHSV Type Strain F1, and were neutralized by the same antiserum, although most of the strains showed low or moderate neutralization titers. None of the isolates were detected by immunofluorescence using a specific monoclonal antibody against a nucleocapsid-related protein of VHSV F1. This is the first report of VHSV isolated from wild Greenland halibut, which represents a new host species for the virus, and it is also the first evidence of VHSV in a location close to the Atlantic coast of North America. This isolation indicates that VHSV is more widely distributed than has been thought, and appears to support a marine origin of this virus.

Advances in the therapy of von Willebrand disease.

von Willebrand disease (vWD) is a very common autosomal inherited bleeding disorder, caused by a quantitative deficiency or a qualitative structural defect of von Willebrand factor (vWF). Two main therapeutic options are available for the treatment of spontaneous bleeding episodes and for prevention of bleeding: desmopressin (DDAVP) and replacement therapy with plasma products. DDAVP is the treatment of choice for most patients with type 1 vWD. In patients with the type 3 disease and in most subjects with type 2 disease, DDAVP alone is ineffective or contraindicated, and it is usually necessary to switch to plasma concentrates containing both factor VIII (FVIII) and vWF. Concentrates subjected to virucidal treatment (e.g. solvent/detergent treatment) during manufacture should always be used in preference to cryoprecipitate. A recombinant vWF concentrate is now undergoing preclinical development and preliminary data suggest it possesses good haemostatic function and may correct the bleeding in vWD after its administration in several animal models. Although treatment of vWD is relatively simple (assuming access to even basic laboratory facilities), actual diagnosis is often far from straightforward, and the patients should be well characterized phenotypically to tailor the treatment to the different types and subtypes of the disease. It is probably wise to refer samples for the characterization to expert laboratories.

Location of superoxide production sites in turbot neutrophils and gilthead seabream acidophilic granulocytes during phagocytosis of glucan particles.

Here we show, by spectrophotometry and enzyme cytochemical methods, that turbot neutrophils and gilthead seabream acidophils responded in a similar way when incubated with PMA or with particulate glucans. Cells stimulated with PMA released high amounts of superoxide both intra- and extracellularly. However, O2- was mainly released intracellularly when cells were incubated with particulate glucans. Small glucan particles were quickly phagocytosed and O2- was initially produced in intracellular vesicles and tubular structures that later fused with the phagosome or with the cell membrane. Large glucan filaments that were not phagocytosed also induced cell stimulation and O2- was also produced in intracellular vesicles which then appeared to fuse with the cell membrane. We conclude that, in stimulated turbot neutrophils and gilthead seabream acidophils, superoxide production is carried out initially in intracellular compartments that are very similar to those described in mammalian neutrophils.

[Pentoxifylline: is it useful in multiple sclerosis?]. / Pentoxifilina: es útil en la esclerosis múltiple ?

INTRODUCTION: Pentoxifylline (PTX) is a phosphodiesterase inhibitor which has been found in studies in vitro to inhibit the production of Th-1 cytokines. It has been postulated that it might be used as a possible coadjuvant treatment for interferon in patients with multiple sclerosis. This would also reduce the potential side effects of interferon. OBJECTIVE. To show the efficacy of PTX in reducing the side effects of interferon, and in the functional improvement of these patients. PATIENTS AND METHODS: We studied 18 patients with remitting-relapsing multiple sclerosis over a period of 18 months; nine patients were given PTX and interferon 800 mg/day simultaneously, and nine patients were treated with interferon alone. The clinical condition was evaluated every three months using the Expanded Disability Status Scale (EDSS) and the Neurological Score (NRS) scales. RESULTS: We found no statistical improvement in the clinical course of EDSS and NRS in either group of patients after treatment for 18 months. The patients treated with PTX have fewer secondary effects due to interferon (fever and myalgia) during the first three months, but these differences between the groups subsequently disappear. In two patients PTX caused transient gastralgias and nauseas. CONCLUSIONS: PTX may be useful as a coadjuvant drug with interferon during the first three months of treatment since some of the side effects of interferon may thus be reduced. However, there seems no justification for using PTX for a longer period since there is no functional improvement.

Ion exchange chromatographic determination of olpadronate, phosphate, phosphite, chloride and methanesulfonic acid.

A method based on single column ion chromatography with UV detection was developed for purity testing and assay of monosodium olpadronate. The analyte aqueous solution is precipitated with methanol to enhance the impurities/olpadronate molar ratio, thus improving purity determination at trace levels. The resulting solution is injected into a standard chromatographic system with UV detector in indirect mode with a Waters IC Pak HR column using diluted nitric acid as the mobile phase. The method was fully validated according to ICH guidelines for the determination of phosphite, phosphate, chloride and methanesulfonic acid in olpadronate being suitable for purity testing and assay.

Pentoxifilina: ¿es útil en la esclerosis múltiple? / Pentoxifylline: is it useful in multiple sclerosis?

Introducción. La pentoxifilina (PTX) es un inhibidor de la fosfodiesterasa que en estudios in vitro inhibe la producción de citocinas Th-1 y se ha postulado como posible tratamiento coadyuvante del interferón (IFN) en los pacientes con esclerosis múltiple, disminuyendo además los posibles efectos secundarios de este fármaco.Objetivos. Comprobar la eficacia de la PTX en la reducción de los efectos secundarios del IFN y en la evolución funcional de los pacientes. Pacientes y métodos. Estudiamos 18 pacientes con esclerosis múltiple remitente-recurrente durante un período de 18 meses; nueve pacientes recibieron simultáneamente 800 mg/día de PTX e IFN y otros nueve enfermos fueron tratados sólo con IFN. Se evaluó su estado clínico cada tres meses aplicando las escalas del estado de incapacidad ampliada (EDSS) y la Neurological Score (NRS). Resultados. No apreciamos mejoría estadísticamente significativa en la evolución de la EDSS y NRS entre ambos grupos de pacientes después de 18 meses de tratamiento. Los pacientes que reciben PTX tienen menos efectos secundarios del IFN (fiebre y mialgias) durante los tres primeros meses, pero estas diferencias entre ambos grupos desaparecen posteriormente. En dos pacientes la PTX produjo transitoriamente gastralgias y náuseas. Conclusiones. La PTX podría ser útil como fármaco coadyuvante del IFN durante los tres primeros meses de tratamiento al reducir algunos efectos secundarios del mismo, pero no parece justificado mantenerlo más tiempo al no existir mejoría del estado funcional (AU)

Molecular characterization and leukocyte distribution of a teleost beta1 integrin molecule.

The beta1 integrin, in combination with the alpha subunit, is responsible for migration of leukocytes into areas of inflammation. Although identified in mammalian species; the beta1 or CD29 molecule has yet to be identified in fish. The present investigation has identified a full-length channel catfish, Ictalurus punctatus, cDNA beta1 molecule composed of 2786 bases and a deduced amino acid sequence of 797 amino acids. The catfish molecule has an amino acid identity ranging from 71.87 to 74.12% with bovine, feline, human, and Xenopus. The channel catfish molecule retains several characteristics of mammalian beta1 molecules, such as four cysteine-rich repeat regions, and eight potential N-linked glycosylation sites. Based on Western blotting the channel catfish beta1 molecule has a molecular mass of approximately 130kDa, essentially the same as that for mammalian species. These results confirm the existence and expression of a beta1 gene in channel catfish, a species phylogenetically distant from mammals.

Identification of a beta 2 (CD18) molecule in a teleost species, Ictalurus punctatus Rafinesque.

Beta 2, in combination with the alpha subunit, is responsible for tight adhesion of leukocytes, especially neutrophils and macrophages, in areas of inflammation. Although identified in mammalian and avian species; the beta 2 or CD18 molecule has yet to be identified in fish. The present investigation has identified a full-length channel catfish, Ictalurus punctatus, cDNA beta 2 molecule composed of 2.8 kb and a deduced amino acid sequence of 772 amino acids. The catfish molecule has an amino acid homology ranging from 54 to 63% with mouse, bovine, rabbit, human and chicken. The channel catfish molecule retains several characteristics of mammalian beta 2 molecules, such as cysteine-rich repeat regions, N-linked glycosylation sites, and several proposed signal sequences. Expression of the beta 2 molecule on the catfish neutrophil cytoplasmic membranes is increased upon phorbol dibutyrate stimulation of the cells. Based on Western blotting and the immunoprecipitation test, the channel catfish beta 2 molecule has a molecular mass of approximately 95 kD, essentially the same as that for mammalian species. However, two additional molecules, perhaps alpha chains, of unexpected molecular mass appear to co-precipitate in the SPIT with the 95 kD CD18 molecule. These results confirm the existence and expression of a beta 2 gene in channel catfish, a species phylogenetically distant from mammalian species.

[Mechanisms of progression of cerebral infarction]. / Mecanismos de progresión del infarto cerebral.

The progression of neurologic symptomatology in the first 48 hours after cerebral infarction appears in more than one third of the patients causing greater morbidity and mortality. Early detection of neurologic deterioration, identification of the predictive factors of worsening and knowledge of the pathogenic mechanisms responsible and their correction make up the basis of the most effective therapy in cerebral infarction. Different hemodynamic alterations (progressive occlusion and growth of the thrombus, insufficiency of colateral circulation, arterial hypotension, etc.) influence, although to different degrees in each patient, in all progressive infarctions. These mechanisms sequentially and progressively condition the necrosis of the areas of ischemic shadow and how new peripheral areas of healthy cerebral tissue are included in the ischemic region, progressively increasing the intensity or the extension of the neurologic manifestations.

Identification of a channel catfish, Ictalurus punctatus (Rafinesque), leukocyte-specific leucine zipper protein.

Five clones isolated from a channel catfish cDNA library were each reactive with monoclonal antibodies (mAbs) C3-1 and 51A only. The size of the cDNA inserts from C3-1 and 51A positive clones was 2.5 Kb and identical based on sequence analysis. Monoclonal antibodies C3-1 and 51A specifically reacted with the expressed product of the 2.5 Kb cDNA clone. The complete DNA sequence indicated that the 2.5 Kb cDNA encoded an approximately 50 Kd protein molecule consisting of 445 amino acids. Sequence analysis showed that this putative protein was a potential leucine-zipper DNA binding protein. Comparison of the deduced amino acid sequence demonstrated homology (14.6 to 19.5%) throughout the sequence of the catfish protein with a group of cytoplasmic-leucine zipper containing proteins of humans; paraneoplastic cellebellar degeneration related (cdr) antigen 2 and 3 with 39.8 to 56.3% homology in the leucine-zipper motif (amino acids 52 through 175 in the catfish protein). This protein was detected in nuclear extracts. cytoplasmic membrane preparations and cytosolic extracts of neutrophils and lymphocytes when reacted with mAbs C3-1 and 51A in an ELISA. However, the intensity of the reactions was dependent upon the cell type and cellular component. The putative cdr protein was not detected with any appreciable intensity in preparations from other cell types. This finding strongly suggests that this protein is expressed in a leukocyte-specific manner and is unique among the cdr group in that it is being expressed in a site that is not immune privileged.

Molecular and functional characterization of channel catfish (Ictalurus punctatus) neutrophil collagenase.

Channel catfish (Ictalurus punctatus) neutrophils, like mammalian neutrophils, contain a variety of enzymes and lytic peptides that participate in pathogen destruction. We have identified and characterized from a channel catfish anterior kidney cDNA library a 1.6 kb cDNA that encodes for channel catfish neutrophil collagenase. The deduced amino acid sequence has a predicted molecular mass of 53 kDa. The putative catfish collagenase has nucleotide and amino acid homology of 51.4% and 45.1%, respectively, with human neutrophil collagenase and 50.4% and 47.1%, respectively, with mouse neutrophil collagenase. Certain regions of the molecule, including the cysteine switch and the putative zinc binding sites, were identical to those in the human and mouse genes. Polyclonal antiserum, prepared to the fusion protein, recognizes proteins from channel catfish neutrophil supernatants with molecular masses of approximately 63, 53 and 28 kDa. Supernatants from phorbol dibutyrate stimulated neutrophils were capable of degrading type I collagen. In addition, the polyclonal antiserum prevented the collagenase activity of the supernatants from stimulated catfish neutrophils; whereas, preimmune serum had no effect on collagenase activity of supernatants. Supernatants from unstimulated cells or the fusion protein did not possess the ability of degrading type I collagen. These results indicate that channel catfish neutrophil collagenases share molecular and functional features with mammalian neutrophil collagenase.

Aggravation of acute ischemic stroke by hyperthermia is related to an excitotoxic mechanism.

OBJECTIVE: To examine whether hyperthermia aggravates cerebral injury in acute ischemia by an excitotoxic mechanism, we studied the relationship between body temperature on admission and CSF concentrations of neuroexcitatory amino acids in 128 patients with acute ischemic stroke of less than 24 h duration. METHODS: Stroke worsening was defined as the percent change between the Canadian Stroke Scale (CSS) at 48 h and the CSS on admission. Infarct volume was measured on days 4-7 on cranial computed tomography. Excitatory amino acids were analyzed using HPLC. RESULTS: Glutamate concentration [median (min.-max.)] was 11 (2-19) micromol/l in hyperthermic patients (body temperature >37.5 degreesC) and 5 (2-22) micromol/l in normothermic patients (p < 0.0001). Glycine concentration in hyperthermic and normothermic patients was 16 (3-21) micromol/l and 9 (3-50) micromol/l, respectively (p < 0.0001). Glutamate was significantly higher in patients with hyperthermia only during the first 12 h after the onset of symptoms. The CSF concentrations of glutamate (r = 0.52; p < 0.0001) and glycine (r = 0.62; p < 0.0001) correlated with body temperature. Body temperature was significantly related to stroke worsening and infarct size, but this effect was dependent on the glutamate effect. CONCLUSION: Glutamate and glycine release during the acute phase of cerebral ischemia could be responsible for the increased brain damage in hyperthermia.

The tropicamide test in patients with dementia of Alzheimer type and frontotemporal dementia.

The tropicamide test was applied to 30 patients with probable Alzheimer's disease (pAD), 12 with frontotemporal dementia (FTD) and 46 healthy subjects. One drop of 0.01% tropicamide was instilled in one eye and one drop of saline solution in the other. The pupil diameter was measured with a Goldmann pupillometer in its basal condition and 10, 15, 20, 25, 30, 35, 45 and 55 minutes afterwards. The results do not show differences between pupil dilation observed in pAD and in FTD; in both groups, from the beginning of the test, the pupil dilated more than in healthy people. A high interindividual variability was observed. The best cutoff point is 38% of interpupillary difference at minute 25 (sensitivity = 63%, specificity = 80%). If we consider the prevalence of AD in a population over 40 years old to be 1.4%, the adjusted positive predictive value of the test would be 4.2%. According to these results, the test is not a true diagnostic marker of AD.

Timing for fever-related brain damage in acute ischemic stroke.

BACKGROUND AND PURPOSE: The association between hyperthermia and early neurological deterioration, increased morbidity, and mortality in acute ischemic stroke is well known. However, the timing at which the cerebral lesion may be aggravated by high temperature has not been firmly established. The aim of this study was to determine the prognostic value of body temperature measured at different times after onset of stroke. METHODS: Axillary temperature was recorded every 2 hour hours for 72 hours in 260 patients with a hemispheric cerebral infarction of <24 hours' duration. A potential infectious focus was examined in all patients with hyperthermia (temperature >37.5 degreesC in any of the assessments). Stroke severity was quantified with the Canadian Stroke Scale on admission. The relationship between the highest temperature recorded in each 6-hour interval from stroke onset and stroke outcome (Canadian Stroke Scale and Barthel Index at 3 months) or infarct volume was evaluated by correlation analyses. The importance of the time at which hyperthermia was first detected was assessed by logistic regression analysis. RESULTS: During the first 72 hours, 158 patients (60.8%) had hyperthermia, and in 57.6% of them an infectious cause was identified. Mortality rate at 3 months was 1% in normothermic patients and 15.8% in hyperthermic patients (P<0.001). The correlation coefficients between the final infarct volume, Canadian Stroke Scale and Barthel Index scores at 3 months, and each temperature recording decreased progressively over time from symptom onset. Hyperthermia initiated within the first 24 hours from stroke onset, but not afterward, was independently related to larger infarct volume (odds ratio [OR]=3.23, 95% CI=1.63 to 6.43; P<0.001) and higher neurological deficit (OR=3.06, 95% CI=1.70 to 5.53; P<0. 001) and dependency (OR=3.41, 95% CI=1.69 to 6.88; P=0.002) at 3 months. The infectious origin of hyperthermia was not associated with poorer outcome or greater infarct volume. CONCLUSIONS: The relationship between brain damage and high temperature is greater the earlier the increase in temperature occurs. However, only body temperature within the first 24 hours from stroke onset is associated with poor outcome and large cerebral infarcts.

[Clinical efficacy of zolmitriptan in migraine]. / Eficacia clínica de zolmitriptán en la migraña.

Zolmitriptan (previously known as 311C90) is a serotoninergic 5-HT1B/D agonist with high oral bioavailability with a double, central and peripheral, action mechanism. Evaluation of its clinical efficacy was developed in a program of clinical studies (search and confirmation of dosis, comparative and long term studies) and through analysis of efficacy in different clinical situations. Zolmitriptan shows a high effectiveness in the treatment of migraine crisis, significantly reduces the headaches by 2 hours of its administration, reduce the symptoms associated with migraine (nausea, photophobia and phonophobia) and improves the quality of life of the migraine patient. The efficacy is independent of the type of migraine characteristics of the patient as well as of the administration of other concomitant medications. The dosis of 2.5 mg of zolmitriptan has been found to be the optimum considering both efficacy and tolerability.

[Detection of soluble interleukin-2 receptor in the serum of patients with non-Hodgkin's lymphoma]. / Determinación del receptor soluble de la interleucina 2 en el suero de pacientes con linfoma no hodgkiniano.

BACKGROUND: Patients with non-Hodgkin's lymphoma (NHL) have increased serum levels of soluble interleukin-2 receptor (sCD25). In this study the authors investigate: a) the value of sCD25, compared to other serum markers, as tumor marker, and b) the relationship of the sCD25 with the response to therapy and prognosis. PATIENTS AND METHODS: Serum interleukin-2 receptor (sCD25) levels were measured at diagnosis in 63 patients with NHL (low-grade lymphoma 30 and high-grade lymphoma 33). RESULTS: High levels of sCD25 were found in these patients compared to a control group (median 1,757 U/ml vs 385 U/ml; p < 0.0001). Significant differences were also found between the high-grade group and the low-grade group, as a whole and within the same Ann Arbor stage. sCD25 showed a correlation coefficient higher than other serum parameters (albumin, LDH, beta 2-microglobulin, uric acid, C-reactive protein) with Ann Arbor stage and with the number of involved lymph nodes or extralymphatic organs. In the high-grade NHL, the median of sCD25 (3,000 U/ml) separates patients with differences in the overall survival (p = 0.0138) and in percentage of complete remisions (p = 0.0079). All the patients with sCD25 < or = 3,000 U/ml reached the remision. The association sCD25 > 3,000 U/ml and albumin < 3.5 g/dl selected to 5 out of 6 patients who failed induction chemotherapy, and only 2 out of 22 who reached the remision. CONCLUSIONS: The sCD25 is the best serum factor for estimating tumor burden in NHL. sCD25 level isolates or associated with albumin provides prognostic information.

[Time for cerebral damage due to hyperglycemia in acute ischemia]. / Tiempo para el daño cerebral por hiperglucemia en la isquemia aguda.

INTRODUCTION: Hyperglycemia increases morbimortality in cerebral infarcts. In animal models, this relationship is only seen during the initial moments of cerebral ischaemia. The time needed in humans for cerebral damage to occur due to hyperglycemia is not known. MATERIAL AND METHODS: We included 194 patients admitted within 24 hours of the start of their first clinical episode of cerebral hemisphere infarction. The glucemia was determined on admission and after 24, 48 and 72 hours. The neurological defect was evaluated on the Canadian scale on the seventh day. The volume of the infarct was determined on a second CT scan done between the fourth and seventh days after the episode. RESULTS: There was a positive association between the volume of the infarct and the glycemias on admission and after 24 hours, but this was not seen in later determinations. The difference in scoring, on the Canadian scale, on the seventh day, between patients with glycemias above or below 120 mg/dl decreased from the time of admission up to the time samples were taken 72 hours later. However, statistical significance persisted during the whole period studied. Nevertheless, in a logistic regression model, the glycemia on admission was the only determination associated with the Canadian scale on the seventh day (OR = 1.02; IC 95% = 1.01-1.02). CONCLUSIONS: There is a clear association between hyperglycemia and the worst stage of the infarct, and this is most intense in the first hours after onset of the clinical features.