Melatonin-induced methylation of the ABCG2/BCRP promoter as a novel mechanism to overcome multidrug resistance in brain tumour stem cells.

BACKGROUND: Current evidence indicates that a stem cell-like sub-population within malignant glioblastomas, that overexpress members of the adenosine triphosphate-binding cassette (ABC) family transporters, is responsible for multidrug resistance and tumour relapse. Eradication of the brain tumour stem cell (BTSC) compartment is therefore essential to achieve a stable and long-lasting remission. METHODS: Melatonin actions were analysed by viability cell assays, flow cytometry, quantitative PCR for mRNA expression, western blot for protein expression and quantitative and qualitative promoter methylation methods. RESULTS: Combinations of melatonin and chemotherapeutic drugs (including temozolomide, current treatment for malignant gliomas) have a synergistic toxic effect on BTSCs and A172 malignant glioma cells. This effect is correlated with a downregulation of the expression and function of the ABC transporter ABCG2/BCRP. Melatonin increased the methylation levels of the ABCG2/BCRP promoter and the effects on ABCG2/BCRP expression and function were prevented by preincubation with a DNA methyltransferase inhibitor. CONCLUSION: Our results point out a possible relationship between the downregulation of ABCG2/BCRP function and the synergistic toxic effect of melatonin and chemotherapeutic drugs. Melatonin could be a promising candidate to overcome multidrug resistance in the treatment of glioblastomas, and thus improve the efficiency of current therapies.

[Pathogenetic bases of epileptogenesis in cerebral cavernomas]. / Bases patogenicas de la epileptogenesis en los cavernomas cerebrales.

INTRODUCTION. Brain cavernoma are a type of arteriovenous malformation that clinically presenting seizures, neurological deficit or bleeding. Hypoxia, neoangiogenesis and metalloproteasas seems to be involved in seizures physiopathology. Our study aims to assess this potential relation by immunohistochemical methods, analyzing hypoxia inducible factor (HIF-1alpha) and metalloproteasa (MMP-9) in tissue surrounding cavernoma. PATIENTS AND METHODS. We selected 17 consecutive cases anatomopathologically diagnosed as cavernoma during 9 years. Immunohistochemical staining was performed for HIF-1alpha and MMP-9. We evaluated the relation between seizures and the scale of uptake of different tissues surrounding cavernoma. RESULTS. Cases with seizures had HIF-1alpha positive uptake in vascular endothelium in 31%, 17% in fibrous tissue and 34% in inflammatory tissue. Besides, it also shows MMP-9 positive uptake in vascular endothelium in 86%, 100% in fibrous tissue and 43% of brain tissue. Statistical analysis by chi-square and odds ratio shows a positive trend towards seizures and the presence of HIF-1alpha and MMP-9 in vascular tissue, fibrous tissue and brain tissue, but no for inflammatory tissue. CONCLUSION. HIF-1alpha and MMP-9, valued by immunohistochemical methods, are related to complications as seizures.

Neuromodulation of the posterolateral hypothalamus for the treatment of chronic refractory cluster headache: Experience in five patients with a modified anatomical target.

INTRODUCTION: Deep brain stimulation (DBS) of the posterior hypothalamus has been found to be effective in the treatment of refractory chronic cluster headache (CCH). METHODS: We report the long-term outcomes of five patients with refractory CCH on whom stimulation of a modified target of approximately 3 mm in radius, which included the posterolateral hypothalamus, the fasciculus mammillotegmentalis, the fasciculus mammillothalamicus and the fasciculus medialis telencephali, was performed. The stereotaxic coordinates were 4 mm from the third ventricle wall, 2 mm from behind the mid-intercommissural point and 5 mm from under the intercommissural line. RESULTS: All patients became pain-free for 1-2 weeks after the procedure, but then needed an average of 54 days to optimize stimulation parameters. After a mean follow-up of 33 months, two remain pain-free, two have an excellent response (>90% decrease in attack frequency) and in one the attacks have been reduced by half. There were no serious adverse events. Permanent myosis and euphoria/well-being feeling were seen in three patients. Other adverse events, such as diplopia, dizziness, global headache of cervical dystonia, were seen transiently related to an increase in stimulation parameters. Attacks reappeared transiently in two patients as a result of cable rupture and when the stimulator was disconnected. CONCLUSIONS: Our results supports the efficacy of DBS in very refractory CCH with a slightly modified hypothalamic target conceived to avoid the lateral ventricle wall so as to extend the stimulated brain area and to decrease the morbidity of potential haemorrhagic complications.

[Review of the target for the deep brain stimulation of chronic cluster headache]. / Revision de la diana para la estimulacion cerebral profunda de la cefalea en racimos cronica.

Cluster headache is included in the group of trigeminal autonomic cephalalgias. Although the pathophysiology of cluster headache has not yet been sufficiently established, the theory of a central origin tells us that this headache is produced by hypothalamic dysfunction. More than 50 patients have been treated with deep brain stimulation of the posterior nucleus of the hypothalamus from 2001. The results show clinical improvement in more than 60% of the cases, opening a promising issue for the treatment of the cluster headache persistent after medical treatment. The surgical target that have been used until now is based on the origin of the cluster headache in the hypothalamic dysfunction. Nevertheless, It has still some open questions as the lack of proving the posterior nucleus of the hypothalamus is the real origin of the cluster headache, the lack of consensus about the anatomy of the surgical target and the variability of the structures stimulated with the surgery. The aim of this article is a review of the target used and propose another surgical target based on physiopathological concepts to explain the improvement with the deep brain stimulation in these patients.

Tratamiento quirúrgico del ictus de la arteria cerebral media / Surgical treatment of the stroke in the middle cerebral artery

Introducción. La craniectomía descompresiva aumenta la supervivencia en los infartos malignos de la arteria cerebral media (ACM). Se analizan los signos radiológicos y clínicos que predicen la evolución maligna del infarto de la ACM,y factores asociados a un peor pronóstico. Pacientes y métodos. Se estudian 30 pacientes divididos en tres grupos: pacientes operados, y pacientes no operados con ingreso en cuidados intensivos o en planta de neurología. La técnica quirúrgica consistióen la creación de una ventana ósea de al menos 10 cm de diámetro y apertura dural. Para la valoración inicial del paciente se utilizó la escala de Glasgow y la escala de ictus del National Institute of Health, y para el seguimiento, la escala modificadade Rankin, el índice de Barthel y la Glasgow Outcome Scale a los seis meses. Resultados. Los pacientes más jóvenes tienen un mejor pronóstico funcional que los mayores de 60 años. La desviación de la línea media mayor de 10 mm se asocia con un peor pronóstico, al igual que volúmenes de tejido infartado mayores de 350 cm3. Menor puntuación en la escala de Glasgow al ingreso se asocia a peor pronóstico vital y a mayor número de secuelas, así como su disminución durante el ingreso. Conclusiones. La edad condiciona la presencia de secuelas en estos pacientes. La presencia de signos clínicos de herniación (anisocoria, menor puntuación inicial o descenso importante en la escala de Glasgow) y radiológicos (desplazamiento de la línea media, volumen infartado) implica un peor pronóstico. La cirugía precoz en aquellos pacientes en que estuviera indicada reduce el número de secuelas y aumenta la supervivencia (AU)

Introduction. Decompressive craniectomy increases the survival rate in cases of malignant middle cerebral artery (MCA) stroke. The imaging and clinical signs that predict a malignant progression of stroke of the MCA are analysed, together with factors associated with a poorer prognosis. Patients and methods. The study involved 30 patients, who were divided into three groups: patients who had undergone surgery, and patients who had not undergone surgery but were admitted to intensive care or to neurology wards. The surgical procedure consisted in creating a bone window with a diameter of at least 10 cm and a dural opening. The initial evaluation of the patient was performed using the Glasgow scale and the National Institute of Health stroke scale; follow-up was carried out using the modified Rankin scale, the Barthel index and the Glasgow Outcome Scale at six months. Results. Younger patients have a better functional prognosis than those over 60 years of age. A deviation of more than 10 mm from the mean line is associated with a poorer prognosis, as are volumes of infarcted tissue above 350 cm3. Lower scores on the Glasgow scale on admission are associated with a poorer prognosis for survival and a higher number of sequelae, as well as their reduction during hospitalisation. Conclusions. Age conditions the presence of sequelae in these patients. The presence of clinical signs of herniation (anisocoria, lower initial score or important drop on the Glasgow scale) and imaging signs (displacement of the mean line, volume of infarcted tissue) imply a poorer prognosis. Early surgery in those patients in whom it is indicated reduces the number of sequelae and increases the rate of survival (AU)

[Surgical treatment of the stroke in the middle cerebral artery]. / Tratamiento quirúrgico del ictus de la arteria cerebral media.

INTRODUCTION: Decompressive craniectomy increases the survival rate in cases of malignant middle cerebral artery (MCA) stroke. The imaging and clinical signs that predict a malignant progression of stroke of the MCA are analysed, together with factors associated with a poorer prognosis. PATIENTS AND METHODS: The study involved 30 patients, who were divided into three groups: patients who had undergone surgery, and patients who had not undergone surgery but were admitted to intensive care or to neurology wards. The surgical procedure consisted in creating a bone window with a diameter of at least 10 cm and a dural opening. The initial evaluation of the patient was performed using the Glasgow scale and the National Institute of Health stroke scale; follow-up was carried out using the modified Rankin scale, the Barthel index and the Glasgow Outcome Scale at six months. RESULTS: Younger patients have a better functional prognosis than those over 60 years of age. A deviation of more than 10 mm from the mean line is associated with a poorer prognosis, as are volumes of infarcted tissue above 350 cm3. Lower scores on the Glasgow scale on admission are associated with a poorer prognosis for survival and a higher number of sequelae, as well as their reduction during hospitalisation. CONCLUSIONS: Age conditions the presence of sequelae in these patients. The presence of clinical signs of herniation (anisocoria, lower initial score or important drop on the Glasgow scale) and imaging signs (displacement of the mean line, volume of infarcted tissue) imply a poorer prognosis. Early surgery in those patients in whom it is indicated reduces the number of sequelae and increases the rate of survival.

[A proposed new target for deep brain stimulation in obsessive-compulsive disorder]. / Propuesta de una nueva diana para la estimulación cerebral profunda en el trastorno obsesivo-compulsivo.

INTRODUCTION: The obsessive-compulsive disorder (OCD) has an incidence in general population of 1.5-3%. If we consider as a positive respond a diminution of the 25-35% in the symptoms of OCD according to the Y-BOCS, and we add the cognitive-behavioral therapy to the pharmacological treatment, only a 40-60% of treated patients would have significant improvement and a 10% of patients with OCD, would be refractory to all type of medical treatment. DEVELOPMENT: Current neurosurgical techniques for resistant cases of OCD interrupt the connections between the frontal lobes and subcortical structures (cingulotomy, capsulotomy). These techniques are ablative and irreversible. It shows the importance of finding a less aggressive technique with better clinical results. Deep brain stimulation (DBS) is an alternative to traditional neurosurgery based in neuromodulation methods. It's considered that the physiopathology of the OCD consists of a dysfunction of the direct and indirect vias that control the extrapiramidal limbic circuit. On the other hand, it had been obtained positive results after DBS of the subthalamic nucleus of three patients with Parkinson's disease and OCD. CONCLUSION: This article has as target the demonstration that bilateral DBS of the limbic part of the subthalamic nucleus is an alternative for the treatment of refractory OCD.

Propuesta de una nueva diana para la estimulación cerebral profunda en el trastorno obsesivo-compulsivo / A proposed new target for deep brain stimulation in obsessive-compulsive disorder

La incidencia del trastorno obsesivo-compulsivo (TOC) en la población general es del 1,5-3%, y secalcula que sólo un 40-60% de los pacientes tratados farmacológicamente tiene mejoría significativa, y un 10% de ellos es refractario a dicho tratamiento. Actualmente, el TOC tiene dos tipos de tratamiento neuroquirúrgico: uno ablativo (cingulotomía,capsulotomía) y otro neuromodulador –estimulación cerebral profunda (ECP)–. Desarrollo. Partiendo del hecho de que la fisiopatología del TOC consiste en una disfunción de las vías directa e indirecta que regulan el circuito límbico extrapiramidal,y de los resultados clínicos de tres pacientes con enfermedad de Parkinson y TOC que, tras recibir ECP en el núcleo subtalámico, han mejorado de las dos patologías, proponemos el área límbica del núcleo subtalámico como diana quirúrgicapara la ECP en el TOC. Las coordenadas estereotáxicas sugeridas serían: x, 8-9 mm lateral a la línea media comisura anterior-comisura posterior; y, 1 mm por delante del punto intermedio comisural; z, 3 mm por debajo de la línea media comisuraanterior-comisura posterior. Conclusión. Este artículo tiene como objetivo demostrar que la ECP bilateral de la parte límbica de los núcleos subtalámicos puede ser una alternativa para el tratamiento del TOC refractario a tratamiento farmacológico

The obsessive-compulsive disorder (OCD) has an incidence in general population of 1.5-3%. If we consider as a positive respond a diminution of the 25-35% in the symptoms of OCD according to the Y-BOCS, and we add thecognitive-behavioral therapy to the pharmacological treatment, only a 40-60% of treated patients would have significant improvement and a 10% of patients with OCD, would be refractory to all type of medical treatment. Development. Current neurosurgical techniques for resistant cases of OCD interrupt the connections between the frontal lobes and subcortical structures (cingulotomy, capsulotomy). These techniques are ablative and irreversible. It shows the importance of finding a less aggressive technique with better clinical results. Deep brain stimulation (DBS) is an alternative to traditional neurosurgery based in neuromodulation methods. It’s considered that the physiopathology of the OCD consists of a dysfunction of the direct and indirect vias that control the extrapiramidal limbic circuit. On the other hand, it had been obtained positiveresults after DBS of the subthalamic nucleus of three patients with Parkinson’s disease and OCD. Conclusion. This article has as target the demonstration that bilateral DBS of the limbic part of the subthalamic nucleus is an alternative for the treatmentof refractory OCD

Complications in subthalamic nucleus stimulation surgery for treatment of Parkinson's disease. Review of 272 procedures.

BACKGROUND: Deep brain stimulation (DBS) is a surgical technique used to alleviate symptoms in patients with advanced Parkinson's disease (PD). It is a reversible procedure and its effect is based on electrical modulation of the nervous system and has considerable advantages in morbidity-mortality when compared to lesion techniques such as thalamotomy and/or pallidotomy. The objective was to evaluate the adverse events during the surgical placement of leads in the subthalamic nucleus for the treatment of Parkinson's disease. METHODS: A retrospective data collection was made in a total of 130 patients in whom we performed 272 procedures for the implant of leads in the subthalamic nucleus between May 1998 and December 2005. All the patients were operated by the same surgeon, in the same institution and with the same surgical methodology. The complications under evaluation were: aborted procedure, misplaced leads, intracranial haemorrhage, seizures, hardware complications and other complications. RESULTS: 130 patients were treated (62 women, 68 men; average age 62 (36-74) years). The average duration of disease from the time of diagnosis to operation was 15.3 years (4-28 years) and the mean follow-up was of 37 months (3-93 months). One hundred and twenty four patients were implanted bilaterally and 6 unilaterally. 62% did not present any complications, 30% had one complication, and 8% more than one complication. Aborted procedures amounted to 5.14% of all procedures, misplaced leads 2.2%, intracranial haemorrhage 3.3%, seizures 4.7%, hardware complications 1.8% and other complications 5.1%. CONCLUSION: Deep brain stimulation surgery is an effective and safe method to treat Parkinson's disease with a low incidence of permanent adverse events.

Effects of the calcium release inhibitor dantrolene and the Ca2+-ATPase inhibitor thapsigargin on spinal nociception in rats.

The effects produced by the intrathecal administration of dantrolene and thapsigargin, measured in several analgesic tests in the rat are described. Dantrolene decreases the release of calcium from intracellular stores and thapsigargin is able to inhibit the reticular Ca2+-ATPase, avoiding intracellular calcium storage. Dantrolene (30-300 nmol/rat) and thapsigargin (3-30 nmol/rat) reduced the nociceptive behavior (biting, scratching, licking; BSL) produced by the NK(1) receptor agonist septide (0.5 microg), without affecting the BSL induced by AMPA (2 microg) or NMDA (4 microg). Also, both drugs elicited analgesia in the tail-flick test but not in the formalin test. The antinociceptive effects induced by thapsigargin were more intense and long-lasting than those produced by dantrolene. These results seem to indicate that the intracellular modulation of calcium homeostasis could be an interesting target in order to induce spinal analgesia.

Effects of intraplantar morphine in the mouse formalin test.

We studied the effects of intraplantar morphine in the formalin test in mice. Intraplantarly administered morphine (30 - 300 microg) induced analgesic effects at lower doses than intraperitoneally administered morphine. Following the administration of [3H]morphine, the % of radioactivity present in brain was the same by either route. In contrast, higher radioactivity values appeared in the injected paw in those mice intraplantarly injected. Since local morphine induces analgesia at doses lower than the intraperitoneally administered drug, especially in the second phase of the test, and the access to brain is undistinguishable, we propose that local morphine enhances central opiate analgesia in the formalin test in mice.

Intrathecal N-methyl-D-aspartate (NMDA) induces paradoxical analgesia in the tail-flick test in rats.

The intrathecal (IT) administration of NMDA in rodents has usually been reported to produce hyperalgesic reactions, although some articles describe that spinal NMDA can lead to analgesia. We show here that the nociceptive behavior (biting, scratching, licking; BSL) observed after NMDA injection (1-8 microg/rat; IT) is followed by a long period of increased tail-flick latencies, not longer detected 24 h after NMDA administration. The NMDA-receptor antagonist CPP (10-100 ng/rat; IT) blocked the BSL behavior induced by NMDA. In the tail-flick test, this antagonist induced analgesia by itself, and was able, at 30 ng/rat, to prevent the NMDA-mediated analgesia. The implication of opiate mechanisms was discarded since naloxone (3 and 10 mg/kg; IP) did not antagonize NMDA-induced analgesia. Finally, the involvement of the intracellular calcium binding protein calmodulin was assessed. The calmodulin inhibitor, calmidazolium (30-300 microg/rat; IT) only blocked the excitatory effect (BSL) without modifying the tail-flick analgesia produced by NMDA (4 microg). These results show that a single intrathecal administration of NMDA sequentially induces both nociceptive and antinociceptive, nonopiate responses in rats.

Comparison of the effects of calmidazolium, morphine and bupivacaine on N-methyl-D-aspartate- and septide-induced nociceptive behaviour.

We have recently shown that spinal calmodulin inhibitors (W-7 and calmidazolium) dose-dependently inhibit the nociceptive reaction (biting, scratching, licking, BSL) evoked by intrathecal N-methyl-D-aspartate (NMDA) and septide, an agonist of the neurokinin (NK) NK1 receptor. To compare this effect with that induced by standard analgesics, we now report a study of the effects of calmidazolium (14420 nmol), bupivacaine (29-582 nmol) and morphine (26-260 nmol) when coadministered intrathecally with either NMDA (4 microg) or septide (0.5 microg). Calmidazolium had the highest potency for inhibiting septide-induced nociceptive behaviour, acting over a dose range of 34-130 nmol (dose eliciting a half-maximal response, ED50, 67 nmol) lower than that of bupivacaine [ED50 234 (115-475) nmol]. Only the highest dose of morphine (260 nmol) inhibited septide-evoked BSL [ED50=133 (69-255) nmol]. Higher doses of morphine could not be tested due to the appearance of an excitatory aversive reaction. Both calmidazolium [ED50=232 (138-388) nmol] and bupivacaine [ED50=123 (59-256) nmol] dose-dependently reduced NMDA-induced BSL reaching an almost maximal inhibition at the highest doses assayed (420 and 291 nmol, respectively). In contrast, morphine had less effect on NMDA-induced behaviour, inducing only a partial reduction of BSL even with the highest dose assayed (260 nmol). Overall, it can be concluded that the calmodulin inhibitor calmidazolium inhibits septide- and NMDA-evoked nociceptive behaviour with a potency and efficacy at least as high as those of morphine and bupivacaine.

[Anomalous response of antibodies in rash-causing viral infections]. / Respuesta anómala de anticuerpos de infecciones virales productoras de rash.

20 pairs of sera from the National System of Seroepidemiological Surveillance of the triple viral vaccine received in the laboratory with febrile rash diagnosis were studied. By using the hemagglutination inhibition test, it was observed an abnormal answer of antibodies to both rubella and measles through a falling of the antibody titre in one or both diseases, or in one of them with seroconversion to the other one. With the aim of defining the existence of a possible polyclonal activation already described in literature, it was decided to study the antibody response to family Herpesviridae (HSV, EBV, CMV, VZV). 80% of answer to these viruses were found. The results are submitted and discussed.

[Report pf the first case of lymphogranuloma venereum (LGV) in an HIV seropositive patient in Cuba]. / Reporte del primer caso de linfogranuloma venéreo (LGV) en un paciente seropositivo para VIH en Cuba.

The most important clinical and epidemiological aspects of the lymphogranuloma venereum as a sexually-transmitted disease are described. We present a summary of the clinical history of an HIV-positive patient who presented with a tumoral lesion in the inguinal region presumptive of lymphogranuloma venereum. The diagnostic value of the polymerase chain reaction (PCR) technique for the establishment of an accurate diagnosis is stressed the epidemiological importance of the report of this sexually-transmitted disease in an HIV-positive patient for the first time in Cuba is also pointed out. A good response was attained with Doxycycline.

[Normalization and application of an ELISA ultramicro for the detection of herpes simplex virus antibodies]. / Normalización y aplicación de un ultramicro ELISA para la detección de anticuerpos al virus herpes simple.

An ELISA ultramicro was normalized for detecting herpes simplex virus antibodies. A study was made on 145 samples from the Pedro Kourí Tropical Medicine Institute and the Blood Bank, both by ELISA ultramicro and immunofluorescence and ELISA, and 98% and 99% of coincidence was reached, respectively. It indicates good correspondence between these techniques.

[Comparison of latex agglutination technics and polyacrylamide gel electrophoresis for the rotavirus diagnosis. Habana City, 1986]. / Comparación de las técnicas de aglutinación por latex y la electroforesis en gel de poliacrilamida para el diagnóstico de rotavirus. Ciudad de la Habana, 1986.

Two methods for the diagnosis of rotavirus are compared with 120 stool samples from children under two years of age who had been hospitalized in Centro Habana Children's Hospital with a clinical diagnosis of acute diarrheal disease and 30 samples of healthy children of the same age group who served as controls. The methods used were latex agglutination (Rotalex, commercially-available kit from ORION DIAGNOSTIC ESPOO, Finland) and polyacrylamide gel electrophoresis. Electronic microscopy was used to define those cases whose results did not coincide with the previous techniques. Comparison was made based on the sensitivity and specificity, cost and easiness of performance. Rotalex was less specific but more sensitive than electrophoresis in polyacrylamide gel. The latter is less expensive, since reactants are available at a lower price in dollars and have no expiration date, is very useful in epidemiologic and outbreak studies, since it detects subgroups of circulating rotavirus and it is a painstaking technique (1-20 samples in 10 hours). Rotalex is simple, rapid (1-20 samples in 30 minutes) and allows individual determinations; therefore, we consider its application in hospitals is feasible.