Therapeutic and diagnostic applications of minor histocompatibility antigen HA-1 and HA-2 disparities in allogeneic hematopoietic stem cell transplantation: a survey of different populations.

Minor histocompatibility antigens (mHags) HA-1 and HA-2 are encoded by biallelic loci, with immunogenic variants, HA-1H and HA-2V, which induce strong HLA-A2-restricted alloreactive T-cell responses, and nonimmunogenic counterparts, HA-1R and HA-2M, which represent functional null alleles that are poorly presented by HLA class I molecules. HA-1 and HA-2 are potential targets of selective graft-versus-leukemia and graft-versus-tumor reactivity after allogeneic hematopoietic stem cell transplantation (HSCT); however, these applications are restricted to a limited number of patients. Here, we show that a far more frequent application of HA-1 and HA-2 disparity relies on their use as markers for the state of host chimerism after allogeneic HSCT. We have determined allelic frequencies of 29.3% and 70.7% for HA-1H and HA-1R, respectively, and of 83.7% and 16.3% for HA-2V and HA-2M, respectively, in >200 healthy individuals from northern Italy. Similar frequencies were observed in nearly 100 patients affected by hematologic malignancies or solid tumors, thus showing that HA-1 and HA-2 variability are not associated with the presence of cancer. On the basis of these data, we predict that HA-1 and HA-2 can be used in 32.8% and 23.5% of Italian transplant patients, respectively, as markers for the state of host chimerism, whereas exploitation of disparity for these mHags for targeted immunotherapy will be possible in 10.7% and 1.1% of Italian patients, respectively. Retrospective HA-2 typing of bone marrow aspirates obtained from a patient during complete remission or recurrence of acute myeloid leukemia after haploidentical HSCT showed the feasibility of using HA-2 as a surrogate marker for disease monitoring. Because of an apparent north-south gradient for HA-1 allelic frequencies, with higher frequencies for the HA-1H variant reported in white populations from Southern Europe as compared with Northern Europe and North America, the diagnostic applicability of HA-1 disparity will be slightly more frequent in transplant patients from the north. Taken together, our data show that determination of HA-1 and HA-2 variability can be an important parameter for the selection of allogeneic stem cell donors, in particular for patients affected by hematologic malignancies without a tumor-specific molecular marker.     

T cell clones expressing the natural killer cell-related p58 receptor molecule display heterogeneity in phenotypic properties and p58 function

A new anti-p58 monoclonal antibody (mAb), termed CH-L, has been used to characterize a minor subset of T lymphocytes co-expressing p58 and CD3 molecules. In two-color immunofluorescence analysis, CH-L⁺CD3⁺ cells represented 0.5 to 6% of the peripheral blood lymphocytes (in 20 healthy donors). Clonal analysis showed that most CD3⁺CH-L⁺ T cell clones expressed the CD8⁺4^? T cell receptor (TcR) α/β⁺ phenotype, while only a few were CD8^?4⁺ TcR α/β⁺, CD8^?4^? TcR α/β⁺ or CD8^?4^? TcR γ/δ⁺. Western blot analysis indicated that the CH-L mAb identifies the same 56-58-kDa diffuse band in both T and natural killer cell (NK) clones. A minority of T cell clones also expressed other NK-related markers such as CD16, CD56 and CD94 and two clones also reacted with the anti-p58 mAb EB6. Interestingly, most clones displayed cytolytic activity in an anti-CD3 mAb-triggered redirected killing assay against the Fcδ receptor⁺ P815 target cells and NK-like activity against K562 and Raji cells. In contrast, the IGROV-1 ovarian carcinoma cell line was resistant to cytolysis by all of these clones. Since p58 molecules have previously been shown to exert regulatory functions on NK-mediated lysis, we investigated whether anti-p58 mAb could also influence cytotoxicity mediated by CD3⁺p58⁺ T lymphocytes. Lysis of P815 target cells, triggered by anti-CD3 mAb, could be inhibited by anti-p58 mAb in 8 out of 12 cytolytic clones tested, while 4 clones were not inhibited. In addition, anti-p58 mAb enhanced the cytolytic activity of 3 clones against IGROV-1 and of 4 other clones against Raji target cells. Taken together, these data indicate that p58⁺ T cells express heterogeneous phenotypes and different forms of TcR and, in most instances, display cytolytic functions. Perhaps more importantly, the p58 molecule appears to modulate the cytolytic activity triggered via the CD3/TcR complex.     

Long-term effects of transdermal and oral estrogens on serum lipids and lipoproteins in postmenopausal women

The transdermal and oral administration of estrogens for one year were compared with respect to the effects on lipid metabolism. Eighty-one postmenopausal women (1.5-3 years after menopause) were randomly divided into three groups. The first two groups received sequential estrogen treatment with either transdermal estradiol (Estraderm TTS, Ciba Geigy; 50 μg/day; 24 women) or 0.625 mg/day conjugated estrogens (Premarin, Wyeth; 20 subjects), respectively. In both groups medroxyprogesterone (10 mg/day per os) was added for 12 days of each cycle. Thirty-five subjects served as control group without therapy. No significant changes in the lipid profile was observed in control subjects after 1 year of follow-up. Serum triglycerides decreased significantly (-10.9 ± 26% S.D.; P < 0.05) in transdermal treated women, whereas it slightly rose in oral estrogen group. Comparable significant decreases in total and low density lipoprotein (LDL) cholesterol (mean range -6.5/-18.0%) were observed in women on estrogen replacement therapy. High density lipoprotein (HDL) cholesterol significantly diminished in transdermal estradiol group, but it rose slightly in the oral estrogen group. Thus the fraction of HDL cholesterol over LDL cholesterol did not change in the transdermal group whereas it significantly rose in subjects treated with oral estrogens. It remains to be established to what extent these differences on lipid metabolism are relevant for the prevention of cardiovascular diseases.     

p53 Immunoreactivity in inflammatory and neoplastic diseases of the uterine cervix

Immunoreactivity for the tumour suppressor gene product p53 is commonly found in many different human malignancies and few premalignant lesions. Data on cervical neoplasms, however, are still lacking. We retrospectively investigated p53 immunoreactivity in 92 lesions of the uterine cervix, including 44 cases of chronic cervicitis, 29 squamous intraepithelial lesions (SILs), and 19 invasive carcinomas. p53 immunoreactivity confined to the basal cell layer, was detected in 74 per cent of cases showing chronic cervicitis and in all cases with low-grade SILs. Conversely, suprabasal and/or diffuse p53 immunoreactivity was exclusively demonstrated in 25 per cent of high-grade SILs and in 74 per cent of invasive carcinomas. The results of this investigation document a high prevalence of p53-immunoreactive malignant tumours of the uterine cervix. In high-grade SILs, p53-immunoreactive cells paralleled the height of involvement by dysplastic changes within the squamous epithelium. A prolonged half-life of the protein is the most likely explanation for the occurrence of p53 immunoreactivity in neoplastic cells. The unexpected finding of p53-immunoreactive cells in inflammatory lesions, though possibly related to an increased proliferation rate of the basal cell compartment, requires further study and underlines the need for a careful approach to p53 immunocytochemistry.     

Surgical treatment of portal hypertension in schistosomiasis

Patients with mansonic schistosomiasis and portal hypertension are usually young, with good liver functional reserve, huge splenomegaly, and hypersplenism detectable only by alterations seen in laboratory tests. Patients who are not operated on do not develop portosystemic encephalopathy (PSE). Angiographic alterations are characteristic and totally different from those observed in patients with liver atrophy or cirrhosis. Digestive tract hemorrhage is the most severe complication. The patient with schistosomiasis is, therefore, a good model for the study of the consequences caused by the surgical treatment of portal hypertension. The evaluation, however, of results published in the literature regarding schistosomiasis is practically impossible. In a review of 130 publications referring to 4,516 surgically treated patients, the absence of prospective or retrospective studies with adequate controls was observed, and only 3.2% of the patients were followed for 5 years. According to available data, a total of 29 different surgical techniques has been employed, 81.6% of which are represented by simple splenectomy, esophagogastric devascularization (EGDS), and splenorenal shunt (SRS). Splenectomy was the surgical procedure showing the highest rate of recurrent hemorrhage (54.3%), and the portocaval shunt had the highest PSE incidence (60%). Portosystemic encephalopathy incidence after SRS was 29%. Thus, in view of a lack of objective information, in 1977 our group started a prospective, randomized study comparing the results of the surgical procedures most widely used in our country: selective portal decompression (SPD), EGDS, and SRS. The randomization was interrupted after 94 patients had been operated on (32 SRS, 30 SPD, and 32 EGDS) because of a highly significant PSE incidence in the SRS group. Preliminary results as of January, 1984, refer to an average follow-up period of 53 months (minimum 29, maximum 77 months). In the SRS group, the late mortality rate was 18.7%, and the incidence of PSE was 31.2%. Among the patients included in this group, 18.7% had arterial hypertension. In the SPD group, late mortality occurred in 10% of the cases, and PSE in 13.3%, while a mild hyperbilirubinemia was seen in 43.3% of patients. Recurrent hemorrhage and ascites had an approximately similar incidence in the 3 groups.

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Resumen Los pacientes con esquistosomiasis mansónica e hipertensión portal generalmente son jóvenes con buena reserva funcional hepática, enorme esplenomegalia e hipersplenismo detectable solamente por las pruebas de laboratorio. Los pacientes nó operados no desarrollan encefalopatia portosistémica (EPS). Los cambios angiográficos son característicos y totalmente diferentes de los que se presentan en pacientes con cirrosis y atrofia hepática. La complicación más grave es la hemorragia digestiva. Por consiguiente, el paciente con esquistosomiasis es un buen modelo para el estudio de las consecuencias que se derivan del tratamiento quirúrgico de la hipertensión portal. Sinembargo, la evaluación de los resultados en la esquistosomiasis que han sido publicados en la literatura es prácticamente imposible. En una revisión de 130 publicaciones relativas a 4516 pacientes operados, se pudo comprobar ausencia de estudios prospectivos o retrospectivos con controles adecuados, y apenas el 3.2% de los casos fué seguido por 5 años. De acuerdo con los datos disponibles, se empleó un total de 29 técnicas quirúrgicas diferentes, 81.6% de los cuales estuvo representado por simple esplenectomía, devascularización esofagogástrica (DEG) y shunt espleno-renal (SER). La esplenectomía fué el procedimiento quirúrgico que tuvo la más alta incidencia de hemorragia recurrente (54.3%) y el shunt portacava la más alta incidencia de EPS (60%). La incidencia de EPS después de SER fué de 29%.Por consiguiente, en vista de la ausencia de información objetiva, nuestro grupo inición en 1977 un estudio prospectivo y aleatorizado para comparar los resultados de los procedimientos quirúrgicos mayormente utilizados en nuestro país: descompresión portal selectiva (DPS), DEG y SER. La aleatorización fué interrumpida después de haber operado los primeros 94 pacientes (32 SER, 30 DPS y 32 DEG) debido a una muy significativa incidencia de la EPS en el grupo con SER. Los resultados preliminares (enero de 1984) se refieren a un período promedio de seguimiento de 53 meses (mínimo 29, máximo 77 meses). En el grupo de SER la mortalidad tardía fué de 18.7% y la incidencia de EPS fué de 31.2%.El 18.7% de los pacientes incluidos en este grupo exhibió hipertensión arterial. En el grupo de DPS la mortalidad tardía fué de 10% y la incidencia de EPS de 13.3%; se observó hiperbilirrubinemia leve en 43.3% de los pacientes. La incidencia de hemorragia recurrente y de ascitis fué aproximadamente igual en los tres grupos.

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Résumé Les malades qui accusent une hypertension portale d'origine bilharzienne sont généralement jeunes. Leurs fonctions hépatiques sont bonnes mais ils présentent une splénomégalie très importante et une hypersplénisme qui se manifeste seulement par des altérations des tests biologiques. Lorsqu'ils ne sont pas opérés ils ne présentent pas de manifestations d'encéphalopathie. Les caractères angiographiques sont très particuliers et totalement différents de ceux qui sont observés chez les sujets dont le foie est atrophié ou cirrhotique. L'hémorragie digestive représente la complication la plus sévère. Le malade atteint de bilharziose hépatique est par conséquent un bon modèle d'étude des conséquences provoquées par le traitement chirurgical de l'hypertension portale. Cependant les résultats du traitement publiés dans la littérature sont ininterprétables. Dans une revue de 130 publications concernant 4516 opérations, l'absence d'études prospectives ou rétrospectives bénéficiant d'un contrôle de qualité a été constatée, 3,2% seulement des malades ayant été suivis pendant 5 ans. En tenant compte des données disponibles 29 techniques chirurgicales ont été employées, 81,6% desquelles sont repésentées par les opérations suivantes: splénectomie, devascularisation et anastomose spléno-rénale. La splénectomie a été suivie du taux le plus élevé de récidive hémorragique (54,3%) et l'anastomose portocave du taux le plus important d'encéphalopathie (60%), l'anastomose spléno-rénale n'étant suivie d'encéphalopathie que dans 29% des cas. En raison du défaut d'information objective notre équipe a entrepris en 1977 une étude prospective faite selon la méthode du choix au hasard des 3 techniques chirurgicales les plus employées au Brésil: l'anastomose spléno-rénale, la décompression portale sélective et la dévascularisation. L'étude fut interrompue alors que 94 malades avaient été opérés (32 anastomose spléno-rénales, 30 décompressions portales sélectives et 32 dévascularisations oesophagogastriques) en raison du taux particulièrement élevé de l'encéphalopathie postopératoire. Les résultats de cette étude menée jusqu'en janvier 1984 répondent à une période postopératoire moyenne de 53 mois (minimum 29, maximum 77 mois). Dans le groupe traité par anastomose spléno-rénale le taux de la mortalité tardive fut de 18,7% et celui de l'encéphalopathie de 31,2%. Parmi les malades de ce groupe, 18,7% présentaient une hypertension artérielle. Dans le groupe traité par décompression portale selective le taux de la mortalité tardive fut de 10%, celui de l'encéphalopathie de 13,3%, cependant qu'une hyperbilirubinémie fut constatée chez 43,3% de ces opères. La récidive hémorragique et l'ascite atteignent un taux très voisin pour les trois opérations.

     

Clearing the clouds: Case‐report and review of the literature

In peritoneal dialysis (PD), a cloudy dialysate is an alarming finding. Bacterial peritonitis is the most common cause, however, atypical infections and non‐infectious causes must be considered. A 46‐year‐old man presented with asthenia, paraesthesia, foamy urine and hypertension. Laboratory testing revealed severe azotaemia, anaemia, hyperkalaemia and nephrotic‐range proteinuria. Haemodialysis was started through a central venous catheter. Later, due to patient preference, a Tenckhoff catheter was inserted. Conversion to PD occurred 3 weeks later, during hospitalization for a presumed central line infection. A month later, the patient was hospitalized for neutropenic fever. He was diagnosed an acute parvovirus infection and was discharged under isoniazid for latent tuberculosis. Four months later, the patient presented with fever and a cloudy effluent. Peritoneal fluid (PF) cytology was suggestive of infectious peritonitis, but the symptoms persisted despite antibiotic therapy. Bacterial and mycological cultures were negative. No neoplastic cells were detected. Mycobacterium tuberculosis eventually grew in PF cultures, despite previous negative molecular tests. Directed therapy was then initiated with excellent response. Thus, facing a cloudy effluent, one must consider multiple aetiologies. Diagnosis of peritoneal tuberculosis is hampered by the lack of highly sensitive and specific exams. Here, diagnosis was only possible due to positive mycobacterial cultures.     

Genetic sperm defects and consanguinity.

BACKGROUND: The existence of a genetic component to human infertility has been suggested, although neither the specific abnormalities involved, nor their genetic mechanism of transmission, are currently defined. We have examined, by transmission electron microscopy (TEM), ejaculate from 1600 males with fertility problems. Among the subjects studied, we focused on a group of patients whose family histories revealed different degrees of consanguinity, in order to evaluate the relationship between consanguinity and particular sperm alterations. METHODS AND RESULTS: A total of 64 consanguineous individuals were identified. In this group, excluding two azoospermic patients, 17 patients (27%) were found to have well recognized genetic ultrastructural defects affecting their entire sperm population: eight subjects had spermatozoa with "stunted tails", four "detached tail" spermatozoa, two "Kartagener's syndrome", two "miniacrosome" and one "round headed" spermatozoa. Since these alterations affect the total sperm population and do not respond to medical treatment, they are suspected of having a genetic origin. The remaining group of 1506 non-consanguineous patients suffered from the same genetic defects in only 15 cases (<1%). CONCLUSIONS: From the data presented, it appears that some very peculiar and rare sperm defects may have a genetic basis since they occur more frequently in consanguineous patients, and are related to different degrees of consanguinity. Since the ejaculate of the remaining patients, both consanguineous and not, showed diverse types of ultrastructural sperm anomalies that did not affect the entire sperm population, they might represent pathologies lacking a genetic basis.     

Binocular interactions and steady-state VEPs. A study in normal and defective binocular vision (Part II)

Background: Recent evidence indicates that an index of binocular activity may be found in some properties of steady-state visual evoked potentials (VEPs), such as amplitude facilitation and phase shortening. We evaluated binocular interactions with steady-state VEPs in normal subjects as well as in patients with concomitant strabismus and defective binocular vision. Methods: Steady-state (8-Hz) VEPs to counterphased sinusoidal gratings (1.2 c/deg spatial frequency) of low contrast (3.2%) were recorded in 19 esotropic patients and in 18 age-matched controls. Patients had either anomalous retinal correspondence (ARC, n =10) or suppression (n=9) in casual seeing conditions (striated glasses). In all subjects, both binocular and monocular VEPs displayed a major component at twice the stimulation frequency (second harmonic), whose amplitude and phase were measured. A binocular interaction index was obtained by comparing binocular VEPs (BVEPs) with the sum (vectorial) of the two monocular VEPs (SMVEPs). Results: In normal subjects, BVEPs were larger in amplitude than SMVEPs (facilitation), and shortened in latency (phase). On average, both ARC and suppression patients displayed loss in amplitude facilitation and absence of phase shortening. However, 50% of ARC patients showed clear VEP facilitation. In both ARC and suppression patients, the amplitude ratio BVEP/SMVEP was negatively correlated with the amount of the angle of deviation. Conclusion: These results suggest that losses in amplitude facilitation and phase shortening of binocular steady-state VEPs reflect abnormal binocular interactions associated with different forms of sensorial adaptation in concomitant strabismus.     

Hiv-1 tat protein suppresses the nerve growth-factor (ngf)-mediated differentiation of PC12 rat pheochromocytoma cell-line

In order to evaluate the effect of the regulatory human immunodeficiency virus-type 1 (HIV-1) Tat protein on the process of neuronal differentiation, two tat-transfected and mock-transfected PC12 cell lines were cultured in the absence or presence of 100-1000 ng/ml of nerve growth factor (NGF). As expected, NGF was able to induce a clearcut morphological differentiation of mock-transfected PC12 into sympathetic-like neurons, also reducing the percentage of cells in S phase. On the other hand, NGF was unable to reduce the percentage of PC12-tat cells in S phase and/or to induce their neuronal differentiation. Only the addition in culture of 5 mu g/ml neutralizing anti-Tat antibody plus 1000 ng/ml NGF was effective in decreasing the percentage of PC12-tat in S phase and inducing partial signs of neuronal differentiation in serum-free cultures. The ability of Tat protein to suppress the neuronal differentiation pathway controlled by NGF further contribute to the definition of its role in tumor promotion during the course of HTV-1 disease.