Gap junctions in human glioblastomas: implications for suicide gene therapy.

Glioblastoma is a very aggressive astrocytic tumor and most patients have 1-year survival time after diagnosis. A promising therapeutic strategy is the local delivery of the herpes simplex virus thymidine kinase gene in the tumor bed followed by ganciclovir treatment. The presence of functional gap junctions is highly relevant for the success of suicide gene therapy. Connexins are expressed in practically all tissues and form gap junctions that allow intercellular communication. Connexin 43 (Cx43) is the major connexin member being expressed in astrocytes but its status in glioblastoma is not well defined. We have investigated by immunofluorescence the presence of Cx43 in 74 human glioblastoma samples; its expression was detected in 77% of the samples analyzed. We report here that glioblastoma is a heterogenous disease as regards Cx43 expression with presentations, in which Cx43 expression is unaltered, reduced or totally lost. A predominant Cx43 cytoplasmic localization was observed in four out of eight primary glioblastoma cultures that we have established. This aberrant localization reduced gap junctionnal intercellular communication by 50 to 75% as compared with primary cell cultures displaying gap junctional plaques. However, the bystander effect evaluated after lentiviral delivery of the herpes simplex virus thymidine kinase gene and ganciclovir treatment was detected in all Cx43-positive primary cell cultures, and it was independant of the Cx43 localization. These findings may have important clinical implications for the design of anticancer cytotoxic therapies that rely on the gap junction-mediated bystander effect for their success.

Sex chromatin in dental pulp: performance of diagnosis test and gold atandard generation / Cromatina sexual en pulpa dental: rendimiento de la prueba diagnóstica y generación del gold standard

The sex chromatin or Barr body is a condensation of chromatin present at the nucleus of cells in female individuals. Their observation is possible in different cell types and is used for the rapid diagnosis of biological sex. The observation of this condensation in cells of the pulp tissue can be useful in the forensic diagnosis of sex, primarily because of the protection it give dental hard tissues, but its diagnostic value has not been tested. The purpose of this study is to analyze the diagnostic value of the observation of sex chromatin in cells of the pulp tissue. We used 40 premolars and third molars 20 from men with a mean age of 20.6 years (SD 8.8) and 20 from women with a mean age of 20.4 years (SD 7.4). The pulp was obtained from teeth extracted and localized 50 cells per plate. The presence of a cell with visible Barr body was considered positive for women. With these results, we evaluated the diagnostic performance of the test. The performance of the test was 100 percent; of the 50 cells observed per plate, mean Barr bodies positive cells was 20.4 (SD 0.44) in female samples. There was no positive cell in preparations of male subjects. The diagnostic test observation of sex chromatin in dental pulp is a reliable test for sex determination. This study provides a gold standard for performance assessment in which the teeth have suffered extreme physical conditions, such as incineration and dumping.

La cromatina sexual o de Barr es una condensación de cromatina presente a nivel del núcleo de las células de individuos de sexo femenino. Su observación es posible en diversos tipos celulares y es utilizado para el diagnóstico rápido del sexo biológico. La observación de esta condensación en células del tejido pulpar puede ser útil en el diagnóstico forense del sexo, principalmente debido a la protección que los tejidos duros dentarios le otorgan, sin embargo su valor diagnóstico no ha sido analizado. El propósito de este estudio es analizar el valor diagnóstico de la observación de la cromatina sexual en células del tejido pulpar. Se utilizaron 40 premolares y terceros molares maxilares, 20 de hombres con una media de edad de 20,6 años (DS 8,8) y 20 de mujeres con una edad media de 20,4 años (DS 7,4). La pulpa fue obtenida a partir de piezas extraídas, se realizó la extirpación pulpar y el tejido fue procesado para H-E. Se seleccionaron aleatoriamente 4 placas histológicas, las cuales fueron observadas de manera sistemática utilizando una microscopia Olimpus CX21 con aumento 100X, localizándose 50 células por placa. La presencia de 1 célula con cromatina sexual visible se consideró positivo para sexo femenino. Con estos resultados se evaluó el rendimiento diagnóstico de la prueba. El rendimiento de la prueba fue del 100 por ciento, de las 50 células observadas por placa, la media de células cromatina sexual positiva fue de 20,4 (DS 0,44) en muestras de sexo femenino. No se observó ninguna célula positiva en preparaciones de sujetos de sexo masculino. La prueba diagnóstica observación de la cromatina sexual en pulpa dentaria es una prueba confiable para la determinación del sexo. Este estudio establece un gold estándar para la evaluación del rendimiento en los que las piezas dentarias han sufrido de condiciones físicas extremas como incineración, inmersión, etc.

Facial tissue thickness in Chilean cadavers with medico-legal / Grosores tisulares faciales en cadáveres de individuos chilenos con fines médicos-legales

The reconstruction of facial features is a key process in human identification with anthropological methods, and represents the ultimate aim of forensic facial reconstruction. This study aimed to record the values of facial tissue thickness of the Chilean population to have a reference table with forensic purposes. We used 40 Chileans cadavers of both sexes, with dates of death between 3 and 48 hours (mean 13.05 hours, SD 10.49) kept in cold storage at -4 º C, aged between 23 and 76 years (mean 45.95 years, SD 11.23), with a BMI average 25.38 (SD 2.27) in men and 26.41 (SD 3.09) in females. We analyzed the facial tissue thicknesses in 20 cephalometric points, eight median and six bilateral paramedian, using the needle puncture--method. We evaluated the presence of sexual dimorphism. The men have a greater thickness at the midpoints, unlike women, where thickness was greater in the paramedian points. However, the differences were significant only in the nasion point. Several studies have reported differences in tissue thickness between men and women, that would justify their systematic determination to develop protocols for facial reconstruction for forensic sculpture. In our study there were no differences, suggesting that different methods of preservation, postmortem time and BMI have influence on gender differences and should be considered in generating population data.

La reconstrucción de la fisonomía facial es una de las claves del proceso de identificación humana con métodos antropológicos y representa el fin último de la reconstrucción con plastía forense. El presente estudio busca registrar los valores de los grosores tisulares faciales de la población chilena para tener una tabla de referencia en el uso con fines médico legales. Se utilizaron 40 cadáveres de individuos chilenos de ambos sexos, con una data de muerte de entre 3 y 48 horas (media 13,05 horas, D.S. 10,49) conservados en cámaras frías a -4º C, con edades entre 23 y 76 años (media 45,95 años, D.S 11,23), IMC promedio hombres 25,38 (DS 2,27) y mujeres 26,41 (DS 3,09). Se analizaron los grosores tisulares faciales en 20 puntos cefalométricos, 8 medianos y 6 paramedianos bilaterales, utilizando el método de punción por agujas. Se evaluó la presencia de dimorfismo sexual. Los hombres presentan un mayor espesor en los puntos medianos, a diferencia de las mujeres en las que grosor fue mayor en los puntos paramedianos. Sin embargo las diferencias sólo fueron significativas en el punto Nasion. Diversos estudios han reportado diferencias en los grosores tisulares entre hombres y mujeres, que justificarían su determinación sistemática al desarrollar protocolos de reconstrucción por plastía forense. En nuestro estudio no se encontraron estas diferencias, lo que sugiere que distintos métodos de conservación, tiempo transcurrido postmortem e índice de masa corporal tiene influencia en las diferencias entre sexos por lo que deben ser consideradas al generar datos poblacionales.

Proceso estiloides elongado: criterios diagnósticos y diagnóstico diferencial: a propósito de un caso / Elongated styloid process: diagnostic criteria and differential diagnosis: a case review

La elongación del proceso estiloides puede presentarse como una entidad patológica en el territorio craneocervical. Existe controversia en relación a los criterios para su diagnóstico, así como sus diagnósticos diferenciales. En el presente reporte se relata el caso de una paciente de 45 años con lumbagos crónicos, cefalea intermitente, dolor cervical y articular agudo, sin historia de quirúrgica o traumática craneocervical. Radiográficamente, destaca la elongación bilateral del proceso estiloides, determinando un diagnóstico presuntivo de Síndrome de Eagle. Se preconiza que los pacientes deben haber sido sometidos a tonsilectomía o trauma para padecer esta patología, condición ausente en este caso. La correcta discriminación de los signos y síntomas permite diferenciar la patología como síndrome estilohioideo. El presente reporte entrega los criterios a considerar en un paciente con el proceso estiloides elongado asociado a patología, con el fin de canalizar esa información hacia un diagnóstico certero.

The elongation of the styloid process may occur as a pathologic entity in the craniocervical area. There is controversy regarding the criteria for its diagnosis and its differential diagnosis. In this report we present the case of a 45 year old female with chronic low back pain, intermittent headache, neck pain and acute articular, no history of surgical or traumatic in the craniocervical area. Radiographically, bilateral elongation of the styloid process were detected, determining a presumptive diagnosis of Eagle syndrome. It requires that patients must have undergone tonsillectomy or trauma to suffer this disease, a condition absent in this case. The correct discrimination of the signs and symptoms distinguish this condition as stylohyoid syndrome. This report delivers the criteria to consider in a patient with elongated styloid process associated with pathology, to use that information and give an accurate diagnosis.

Rol de las características clínicas e histológicas como factores pronósticos para la sobrevida en pacientes con carcinoma de células escamosas de la cavidad oral / Oral squamous cell carcinoma: restrospective analysis of 36 cases

Background: Survival of patients with oral squamous cell carcinoma is low and depends mostly on TNM staging of the tumor. Aim: To perform a retrospective analysis of a series of patients with oral squamous cell carcinoma. Material and Methods: Retrospective review of patients with the diagnosis, seen between 1995 and 2006 in a regional hospital. Host inflammatory response and pattern of tumor invasion were assessed using the staging proposed by Bryne et al. Results: The medical records of 36 patients aged 39 to 89 years, were reviewed. During the study period, 15 patients died. Better survival was associated to a low pattern of tumor invasion and a high inflammatory response and the topographic location of the tumor. Conclusions: Inflammatory response, tumor invasion and location are associated with survival in oral squamous cell carcinoma.

Introducción: El carcinoma de células escamosas de la cavidad oral (CCECO) es una patología cuyo comportamiento es producto de interrelaciones con el huésped, esto es, por el patrón de invasión (PI) histopatológica y la respuesta inflamatoria (RI). El objetivo de este estudio es analizar las características clínicas e histopatológicas como factores pronóstico, en términos de supervivencia (SV) en pacientes con CCECO. Material y Método: Serie de casos. Se incluyeron pacientes diagnosticados en el Hospital Regional de Talca y Hospital Base de Curicó entre los años 1995 y 2006. Se revisaron las fichas clínicas y biopsias de 36 pacientes con CCECO. Se determinó el Frente Invasivo Tumoral (FIT), evaluándose los parámetros propuestos por el sistema de graduación de Bryne (PI y RI) y factores de importancia clínica como localización topográfica de la lesión, edad y género, relacionándolos con SV mediante curvas de Kaplan-Meier y Log Rank test. Posteriormente, se aplicó una regresión de Cox para obtener un análisis multivariado y cálculo de RR. Del total de casos del estudio, 15 pacientes fallecieron por CCECO. Resultados: La mayor SV se asoció a un bajo escore de PI y una alta RI respectivamente (RR 1,5). La localización topográfica de la lesión se relacionó significativamente con la SV, no así el grupo de edad. Conclusiones: Nuestros resultados sugieren que la ubicación de la lesión es un factor importante en el pronóstico de la enfermedad y que una respuesta inmune/inflamatoria adecuada, expresada en un bajo escore de RI, mejora el pronóstico de SV en pacientes con CCECO.

Nanoodontología: el futuro de la odontología basada en sistemas nanotecnológicos / Nanodentistry: the future of dentistry based on nanotechnology systems

El desarrollo científico sin precedentes alcanzado en los últimos años por la nanotecnología ha generado una nueva dimensión en el desarrollo tecnológico, médico y económico que ha marcado un impacto global. Los avances en estrategias de diagnóstico, tratamiento, y prevención de las enfermedades preveen un importante rol de la nanotecnología en el emergente campo aplicado a la odontología denominado nanoodontología. Entre sus principales desafíos destaca el alcanzar una mejor comprensión de las bases fisiopatológicas de las enfermedades, nuevas estrategias de diagnóstico, y el desarrollo de terapias altamente efectivas. La nanotecnología indica ser el camino de la evolución, y el desarrollo de la odontología del siglo 21. Este artículo, presenta las tendencias, y desafíos de nanotecnología aplicada a la odontología, provee de una visión global del desarrollo de la nanoodontología y su impacto futuro.

The unprecedented scientific development achieved in recent years by the nanotechnology has created a new dimension in the technological, medical and economic development set a global impact. Advances in strategies for diagnosis, treatment, and prevention of diseases foresee an important role of the nanotechnology in the emerging field applied to dentistry called nanodentistry. Among its highlights the main challenges to achieve a better understanding of the pathophysiological bases of diseases, new diagnostic strategies, and development of highly effective therapies. Nanotechnology shows to be the path of evolution, and development of dentistry in the 21st century. This article presents the trends and challenges of nanotechnology applied to dentistry, provides an overview of nanodentistry development and its future impact.

Anatomía quirúrgica del drenaje venoso en la región del triángulo carotídeo / Surgical anatomy of jugular vein tributaries

Background: The main tributaries of the internal jugular vein join at cervical level through the venous thyrolinguofacial trunk. This trunk is classically described as formed by the union of the facial, superior thyroid and lingual veins. Aim: To evaluate variations in the formation of the thyrolinguofacial trunk in human cadavers. Material and Methods: Thirty hemi-necks were dissected in human cadavers of nine men and six women, with ages ranging between 29 and 50 years, and analyzed by direct observation. Results: In 16 hemi-necks (53.3 percent), the trunk was thyrolinguofacial; in seven (23.3 percent), it was linguofacial; in six (20 percent), it was thyrolingual and in one case (3.3 percent) it was thyrolinguo pharyngofacial. No thyrofacial trunk formation was found. There was a correlation between the diameter of the internal jugular vein and of the venous trunk formed by these veins. Conclusions: There are morphological changes in the formation patterns of facial, lingual and superior thyroid veins. This information is useful for surgical oncology, plastic surgery, head and neck surgery and radiology.

Los principales afluentes de la vena yugular interna se incorporan a nivel cervical por medio del tronco venoso tirolinguofacial. Clásicamente se ha descrito su formación por la unión de la vena facial, lingual y tiroidea superior. El objetivo de este estudio fue determinar las variaciones en la presentación del tronco tirolinguofacial en cadáveres humanos de la región del Maule, Chile. Se disecaron 30 hemicuellos pertenecientes a 15 cadáveres humanos de ambos sexos, con un rango de edad entre 29 y 50 años, y se analizaron mediante observación directa. Se observó la formación de tronco venoso en la totalidad de los casos, siendo 53,3 por ciento correspondiente al tronco de tipo tirolinguofacial, 23,3 por ciento linguofacial, 20 por ciento tirolingual y sólo un 3,3 por ciento el tronco tirolinguofaringofacial. No se encontró la formación del tronco venoso de tipo tirofacial. Además se encontró una correlación significativa entre el diámetro de la vena yugular interna y el tronco venoso conformado por estas venas. Por lo tanto, existen variaciones morfológicas en los patrones de conformación de las venas facial, lingual y tiroidea superior, siendo estos datos de importancia para áreas de cirugía oncológica, cirugía plástica, cirugía de cabeza y cuello y radiología.

Schwannoma maligno en la mandíbula: reporte de un caso / Malignant Schwannoma in mandible: a case report

El schwannoma maligno es una neoplasia cuyo origen está dado por tejido nervioso periférico. Rara vez se presenta en cabeza y cuello, más escasamente se encuentra en el territorio maxilofacial. A continuación se reporta desde el punto de vista radiológico el caso de un paciente de sexo masculino, de 9 años de edad, que presentó un aumento de volumen de crecimiento rápido, expansivo, ubicado en el cuerpo y rama mandibular derecha con diagnóstico de neurosarcoma maligno de mandíbula o schwannoma maligno mandibular con compromiso del canal mandibular, canal y foramen mentoniano, tejido blando y tejido óseo del sector.

The malignant schwannoma is a neoplasia whose origin is given by peripheral nerve tissue. It rarely appears in the head and neck, and even less frequently in maxillofacial territory. Here is a case report of a male patient, 9 years old, with a rapidly growing mass, expansive growth, located in the body and right mandibular ramus diagnosed as a malignant neurosarcoma or malignant schwannoma of the mandible, with mandibular canal, mental canal and foramen, soft tissue and bone involvement of the region.

Structure of the human 3alpha-hydroxysteroid dehydrogenase type 3 in complex with testosterone and NADP at 1.25-A resolution.

The first crystallographic structure of human type 3 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD3, AKR1C2), an enzyme playing a critical role in steroid hormone metabolism, has been determined in complex with testosterone and NADP at 1.25-A resolution. The enzyme's 17beta-HSD activity was studied in comparison with its 3alpha-HSD activity. The enzyme catalyzes the inactivation of dihydrotestosterone into 5alpha-androstane-3alpha,17beta-diol (3alpha-diol) as well as the transformation of androstenedione into testosterone. Using our homogeneous and highly active enzyme preparation, we have obtained 150-fold higher 3alpha-HSD specificity as compared with the former reports in the literature. Although the rat and the human 3alpha-HSDs share 81% sequence homology, our structure reveals significantly different geometries of the active sites. Substitution of the Ser(222) by a histidine in the human enzyme may compel the steroid to adopt a different binding to that previously described for the rat (Bennett, M. J., Albert, R. H., Jez, J. M., Ma, H., Penning, T. M., and Lewis, M. (1997) Structure 5, 799-T812). Furthermore, we showed that the affinity for the cofactor is higher in the human 3alpha-HSD3 than the rat enzyme due to the presence of additional hydrogen bonds on the adenine moiety and that the cofactor is present under its reduced form in the active site in our preparation.

Crystallization and preliminary X-ray crystallographic analysis of the human type 3 3 alpha-hydroxysteroid dehydrogenase at 1.8 A resolution.

In androgen-sensitive target tissues, 3 alpha-hydroxysteroid dehydrogenase regulates the androgen receptor (AR) activity by catalyzing the inactivation of 5 alpha-dihydrotestosterone (the most natural potent androgen) to 5 alpha-androstane-3 alpha,17 beta-diol. In this report, the crystallization of a human prostatic type 3 3 alpha-hydroxysteroid dehydrogenase, a member of the aldo-keto reductase superfamily, is described. Two different crystal forms of the complex between the human type 3 3 alpha-HSD, NADP(+) and testosterone have been obtained using PEG as precipitant. Crystal form I, which diffracts to 1.6 A, belongs to the monoclinic space group P2(1), with unit-cell parameters a = 55.07, b = 87.15, c = 76.88 A, beta = 107.37 degrees and two subunits in the asymmetric unit. A complete data set has been collected at 1.8 A. Crystal form II, which diffracts to 2.6 A, belongs to the rhombohedral space group R32, with unit-cell parameters a = b = 143.59, c = 205.86 A, alpha = beta = 90, gamma = 120 degrees and two subunits in the asymmetric unit.

Asymmetric synthesis of functionalized carbocycles and heterocycles.

Anions of 1-halo-4-hexenyl phosphonamides derived from chiral, enantiopure C2 symmetrical 1,2-diamino cyclohexane react at the gamma-position in conjugate addition reactions with alpha, beta-unsaturated carbonyl compounds such as cyclopentenone, 4-(H)-furanone, pyrroline-2-one, and cinnamates to give functionalized adducts. Addition to imines is also possible. The adducts can be transformed into enantiopure or enriched carbocyclic and heterocyclic compounds bearing useable functionality.

Familial intracranial aneurysms: recurrence risk and accidental aggregation study.

BACKGROUND: The Saguenay-Lac-Saint-Jean (SLSJ) region is a geographically isolated area (population 285,955) located in the Northeastern part of the Province of Quebec, Canada. Using a population-based register, the genealogical reconstruction of 502 individuals with ruptured intracranial aneurysm (RIA) showed a familial aggregation (the presence of aneurysm in two or more first- to third-degree relatives) for 144 (28.7%) of them; this proportion is much higher than reported elsewhere. OBJECTIVE: In order to assess the genetic predisposition to RIA in the SLSJ population, the objective of the present study is to compare familial and non-familial cases and to provide an estimate of the recurrence risk ratio for siblings. RESULTS: The age at the time of rupture, the number of intracranial aneurysms for each patient and the location of RIAs were not statistically different in the familial versus the non-familial group. Of the 3449 siblings, 20 (0.58%) had suffered a RIA. The recurrence risk ratio calculated for siblings (defined as the risk of disease among siblings divided by the estimated population prevalence) is 1.6 (CI 95% 1.0-2.4). In other respects, we observed very large kinships in the SLSJ population, with an average number of siblings of 7.2 (SD +/- 3.4), ranging from 0 to 17 individuals. With such large families and on the basis of chance alone, we expected 31.3% of the patients to have at least one first- to third-degree relative with RIA. CONCLUSION: These data show that siblings of patients with RIA in the SLSJ population have a greater risk of RIA than the general population. Nevertheless, the largest part of the familial occurrence observed in the SLSJ region can be explained by accidental aggregation, due to large kinships. We propose that, in this population, an underlying genetic predisposition must be suspected only when three or more cases of RIA are identified among first- to third-degree relatives.

Epidemiological study of reptured intracranial aneurysms in the Saguenay-Lac-Saint-Jean region (Quebec, Canada).

BACKGROUND: Using a population-based register of the Saguenay-Lac-Saint-Jean region (Quebec, Canada), the genealogical reconstruction of 533 individuals with intracranial aneurysm (IA) showed a familial aggregation (the presence of aneurysm in two or more first- to third-degree relatives) for 159 (29.8%) of them; this proportion is much higher than reported elsewhere. OBJECTIVE: As part of an ongoing project to assess a genetic predisposition to intracranial aneurysms in the Saguenay-Lac-Saint-Jean population, the objective of the present study was to determine whether age-specific rates of reputed cerebral aneurysms were higher than in other populations. DESIGN: A retrospective study of cases of proven ruptured IAs which were hospitalized during the 1973 to 1992 period was conducted. Age-adjusted rates were computed and compared to those reported in the Helsinki population. RESULTS: We identified 412 cases of ruptured aneurysms. The age-adjusted incidence rate was 7.2/100,000/year (6.2 for men, 8.1 for women), which is similar to the incidence rates reported in other studies. Although the mean age at time of rupture was younger (46.6 years +/- 13.8) than usually reported, no increase in age-specific incidence rates was detected. CONCLUSIONS: The results of this epidemiological study neither support nor reject the hypothesis of a genetic predisposition to intracranial aneurysms in the Saguenay-Lac-Saint-Jean population.

Identification of stable plant cystatin/nematode proteinase complexes using mildly denaturing gelatin/polyacrylamide gel electrophoresis.

The biochemical interactions between two cystatins from rice seeds, oryzacystatin I (OCI) and oryzacystatin II (OCII), and the cysteine proteinases from three plant parasitic nematodes, Meloidogyne hapla, M. incognita and M. javanica, were assessed using standard protease assays and mildly denaturing gelatin/polyacrylamide gel electrophoresis (gelatin/PAGE). Activity detected in extracts of preparasitic second-stage larvae (J2) from M. hapla was optimal at pH 5.5 and was inhibited in vitro by the cysteine proteinase inhibitors trans-epoxysuccinyl-L-leucylamido-(4-guanidino) butane, hen egg cystatin, OCI, and OCII. As demonstrated by class-specific activity staining, all the activity measured between pH 3.5 and pH 7.5 was accounted for by a major proteinase form, Mhp1, and two minor forms, Mhp2 and Mhp3. Mhps were also detected in extracts and excretions of parasitic J2 and adult females, indicating their continuous expression throughout development of M. hapla, and their possible involvement in the extracellular degradation of proteins. Interestingly, the two plant cysteine proteinase inhibitors OCI and OCII showed different degrees of affinity for the major proteinase form, Mhp1. Both inhibitors almost completely inactivated this proteinase in native conditions but, unlike OCII, OCI conserved a high affinity for Mhp1 during mildly denaturing gelatin/PAGE, showing the differential stabilities of the OCI/Mhp1 and OCII/Mhp1 complexes. In contrast to Mhp1, the major cysteine proteinases detected in the two closely related species M. incognita and M. javanica were strongly inhibited by OCII, while the inhibition of OCI was partly prevented during electrophoresis. This species-related efficiency of plant cystatins against nematode cysteine proteinases could have practical implications when planning their use to control nematodes of the genus Meloidogyne.

A study of inbreeding and kinship in intracranial aneurysms in the Saguenay Lac-Saint-Jean region (Quebec, Canada).

The genealogies of 533 individuals with an intracranial aneurysm (IA) born in the Saguenay-Lac-Saint-Jean region, a geographically isolated area located in northeastern Quebec, were reconstructed using a population-based register. A control group consisting of three individuals of the same sex and born on the same day and in the same municipality than the IA patients was created; the genealogies of the 1599 controls were also reconstructed. The coefficients of inbreeding and kinship were calculated. Familial aggregation, i.e. the presence of IA in two or more first- to third-degree relatives, was also sought. The mean inbreeding coefficient was lower in the IA group than in the control group (7.92 x 10(-4) versus 10.04 x 10(-4)). The mean kinship coefficient was higher in the IA group than in the control group (2.17 x 10(-4) versus 1.55 x 10(-4)). Forty-eight IA patients (9.0%) were first-degree relatives compared to only 1.9% of the control individuals. The proportion of individuals showing familial aggregation was higher in the IA group than in the control group (29.8% and 18.6% respectively). These results strongly suggest that some IA are genetically determined in this population.

Assessing the stability of cystatin/cysteine proteinase complexes using mildly-denaturing gelatin-polyacrylamide gel electrophoresis.

A method for assessing the stability of cystatin/cysteine proteinase complexes using mildly-denaturing gelatin-polyacrylamide gel electrophoresis (gelatin-PAGE) is described. As suggested by the use of well-known cystatins (human stefins A and B, and oryzacystatins I and II) and the plant cysteine proteinase papain, the ability of cystatin/cysteine proteinase complexes to remain stable during electrophoresis is associated with the degree of affinity between the enzyme and the inhibitor (and inversely associated with the Ki values), at least with the disulfide bond-lacking cystatins. Complexes with Ki values > or = 10(-8) M (weak interactions) are partly or completely dissociated under the conditions used, while those with lower Ki values (strong interactions) remain stable. As shown by the differential effects of two plant cystatins, oryzacystatins I and II, against a cysteine proteinase present in crude (complex) extracts from a plant pest -- the two-spotted spider mite (Tetranychus urticae Koch), the gelatin-PAGE procedure is suitable for studying the ability of cystatins to form highly stable complexes with cysteine proteinases, without the need for prior purification steps. Considering the well-recognized potential of proteinase inhibitors for pest and pathogen control, this analytical approach will be useful for rapidly assessing the respective potential of various cystatins for protection of plants, animals, and humans.