Keratin immunohistochemistry in renal cell carcinoma subtypes and renal oncocytomas: a systematic analysis of 233 tumors

Keratin immunohistochemistry represents a widely applied differential diagnostic tool in surgical pathology. To investigate the value of keratin subtyping for the diagnosis among histological subtypes of renal cell carcinoma and oncocytomas, we performed a detailed immunohistochemical study, applying 22 different monoclonal keratin antibodies on a large series of 233 renal tumors [125 conventional, 22 chromophobe, and 20 papillary (12 type-1, 8 type-2 tumors) cancers and 66 oncocytomas] using a tissue microarray technique. Immunoreactivity for keratin 7, 8, 18, and 19 was present in all tumor entities, albeit in varying quantities. With antibodies directed against keratins 8 and 18, oncocytomas showed a distinct perinuclear and punctate dot-like pattern, which was not observed in renal cancer specimens. The only tumors showing immunoreactivity for keratin 20 were two type-2 papillary cancers. All other monospecific keratin antibodies yielded consistently negative results. Overall, in contrast to some recent publications, keratin subtyping generally appeared to be of additional value only for the differentiation of renal epithelial tumors. Hence, with respect to differential diagnostic value, Hales colloidal iron stain and vimentin immunostaining are still the most useful tools in renal tumor pathology.     

Type I interferon gene transfer sensitizes melanoma cells to apoptosis via a target activity on mitochondrial function

Our previous article reported that retroviral transduction of human type I consensus interferon-coding sequence into two human melanoma cells increased their susceptibility to cisplatin-induced apoptosis. Importantly, primary melanoma cells were significantly more sensitive to cisplatin-induced apoptosis with respect to metastatic melanoma cells. The aim of this study was to elucidate the subcellular mechanisms involved in this interferon-induced apoptotic proneness. Our results indicate that 1) cisplatin-induced apoptosis can be referred to as the type II apoptosis, ie, to the mitochondrially driven cascade; 2) treatment of interferon-producing melanoma cells with other type II apoptotic stimuli, such as radiation or staurosporine, also resulted in massive apoptosis, whereas type I stimuli, ie, anti-Fas, were ineffective; 3) interferon sensitization involved the caspase cascade in primary melanoma cells and the alternative pathway represented by cathepsin-mediated apoptosis in metastatic melanoma cells; 4) interferon production sensitizes cells to apoptosis by inducing, as the earliest event, mitochondrial membrane hyperpolarization. These results suggest that constitutive production of type I interferon by melanoma cells can act as an intracellular booster capable of increasing cell proneness to apoptosis by specifically modifying mitochondrial homeostasis and independently from the apoptotic cascade involved.     

Prophylactic and therapeutic efficacy of antibodies to a capsular polysaccharide shared among vancomycin-sensitive and -resistant enterococci

Enterococci are important nosocomial pathogens that are increasingly difficult to treat due to intrinsic and acquired resistance to antibiotics, including vancomycin. A recently described capsular polysaccharide (CP) isolated from Enterococcus faecalis 12030 was used to evaluate the potential efficacy of active or passive immunotherapy regimens as adjunctive treatments. Evaluation of protective efficacy was carried out in immunocompetent mice challenged intravenously (i.v.) with live enterococci. In nonimmune mice, i.v. inoculations resulted in high levels of bacteria in kidneys, spleens, and livers 5 days after challenge. Mice immunized with four 10-microg doses of CP antigen/mouse were protected against challenge with the homologous E. faecalis strain. High-titer opsonic immunoglobulin G was also induced by immunizing rabbits with the purified CP, and passive transfer of this antiserum to mice produced significantly lower bacterial counts in organs than did normal rabbit serum or sterile saline. Antibodies to the polysaccharide isolated from E. faecalis 12030 were protective against Enterococcus faecalis OG1RF and against two serologically related, vancomycin-resistant Enterococcus faecium clinical isolates. Antibodies to this CP antigen were also effective as a therapeutic reagent in mice when passive therapy was initiated 48 h after live bacterial challenge. These data indicate that CP antigens from enterococci are potential targets of protective antibodies and that these antibodies may be useful for prophylaxis and treatment of enterococcal infections.     

In vitro and in vivo modulation of MDR1/P-glycoprotein in HIV-infected patients administered highly active antiretroviral therapy and liposomal doxorubicin

P-glycoprotein (P-gp) transports a wide range of structurally unrelated drugs, such as HIV protease inhibitors (PIs) and cytotoxic compounds such as anthracyclines. Because modification of P-gp phenotype and function is an important underlying mechanism of drug interactions, the current study was conducted in order to evaluate whether highly active antiretroviral therapy (HAART), HIV plasma viral load (VL), or cancer chemotherapy may induce in vivo changes of P-gp phenotype in peripheral blood mononuclear cells (PBMCs) from HIV-infected treatment-naive and -experienced subjects at different stages of HIV infection and/or disease, including patients with HIV-associated Kaposi sarcoma (KS). Our results show that neither HAART nor HIV VL, nor the stage of HIV infection and/or disease, significantly alter P-gp expression on PBMCs. In particular, surface P-gp expression is expressed at low levels by T-cell subsets, B cells, and NK cells, whereas almost all monocytes are double positive and these results are not modified by HIV PI-containing regimens. By contrast, a significant phenotype modification is detected in PBMCs from AIDS/KS patients after challenge with the liposomal formulation of the anthracycline doxorubicin (L-DOX) with the higher expression reached 24 hours after the end of the drug infusion. In addition, accumulation of L-DOX is unaffected by P-gp-mediated drug efflux as documented by in vitro experiments, in sharp contrast to the kinetic of free DOX, based on HIV PI blockade experiments. Finally, P-gp expression was found in KS spindle cells from HIV-infected treatment-naive AIDS/KS patients. We conclude that P-gp phenotype in PBMCs and specific subsets is not altered by HAART and/or HIV, whereas a significant increase is induced by specific anticancer drugs such as L-DOX. Moreover, HIV PIs possess an inhibitory effect on P-gp function that may improve DOX sensitivity in KS spindle cells.     

Immunogenic properties of Pseudomonas aeruginosa mucoid exopolysaccharide.

Previous studies have shown that antibodies to Pseudomonas aeruginosa mucoid exopolysaccharide (MEP) can be divided into two types on the basis of their functional activity. One type is able to mediate opsonic killing in conjunction with leukocytes and complement, and the other type is not. We investigated, in mice, the properties of this antigen associated with elicitation of opsonic killing antibody. We found that smaller-sized material (Kav = 0.26), which has been tested as a human vaccine, elicited opsonic killing antibody in mice at low doses (1 to 10 micrograms) and at doses of greater than or equal to 40 micrograms, only nonopsonic killing antibody was produced. A similar dose effect was seen with heat-killed mucoid P. aeruginosa cells. After immunization with high doses of this MEP or the heat-killed cells, the mice were refractory to induction of opsonic killing antibody no matter what dose of MEP was used as a booster. In contrast, a larger-molecular-weight preparation (Kav = 0.05) elicited opsonic killing antibody over a wide dose range (1 to 400 micrograms). Additionally, 50 micrograms of the larger-sized preparation could overcome the suppression induced by 50 micrograms of the smaller material. Suppression elicited by 50 micrograms of the smaller material could be adoptively transferred to nonimmune mice with the T-cell fraction of spleen cells. These results indicate that both molecular size and dose are critical determinants for eliciting opsonic killing antibody to mucoid P. aeruginosa after immunizing with MEP.     

Effects of sodium fusidate in animal models of insulin-dependent diabetes mellitus and septic shock.

We have evaluated the effects of the novel immunosuppressant sodium fusidate (fusidin) in the non-obese diabetic (NOD) mouse and in D-galactosamine (D-Gal)-presensitized BALB/c mice challenged with the bacterial superantigen, Staphylococcus aureus enterotoxin B (SEB) or with the endotoxin, Escherichia coli lipopolysaccharide (LPS). The NOD mouse model has clinical and histoimmunological features similar to those of human insulin-dependent diabetes mellitus (IDDM). The SEB- and LPS-treated BALB/c mouse models exhibit pathogenic similarities with human septic shock conditions. In the NOD mouse, fusidin suppressed the spontaneous development of insulitis (mean inhibition 73%) and hyperglycaemia (IDDM incidence 25% versus 0%) when administered at 40 mg/kg five times weekly for 8 consecutive weeks from the fourth week of age; concurrently treated animals exhibited reduced percentages of splenic T lymphocytes. This anti-diabetogenic effect was confirmed in the accelerated model of diabetes induced in the NOD mouse with cyclophosphamide (CY) (IDDM incidence 55% versus 21-6% using dosages of fusidin from 40 to 80 mg/kg five times weekly); protection from IDDM development was achieved even when the drug (80 mg/kg/day) was first administered 7 days after CY challenge. In contrast, fusidin did not reverse hyperglycaemia when administered to CY-treated animals within 3 days of IDDM development. In the two models of septic shock, prophylactic treatment with fusidin, 80 mg/kg given three times for 2 days prior to D-Gal/SEB or D-Gal/LPS challenge, drastically reduced the lethality compared with D-Gal/buffer-treated mice. This effect may depend on the inhibitory action of fusidin on the secretion of cytokines such as interferon-gamma and tumour necrosis factor-alpha, the serum levels of which were greatly diminished in the fusidin-treated mice (mean inhibition 50-90%). These results demonstrate that fusidin may have a role in the treatment of cell-mediated autoimmune diseases and cytokine-mediated infectious diseases in humans.     

Effectiveness of combined GnRH analogue plus raloxifene administration in the treatment of uterine leiomyomas: a prospective,randomized, single-blind,placebo-controlled clinical trial

BACKGROUND: Raloxifene hydrochloride is a synthetic non-steroidal drug used for the prevention and treatment of post-menopausal osteoporosis. Pre-clinical and clinical data have shown that raloxifene may have a beneficial effect on leiomyomas. The aim of this prospective single-blind, randomized, placebo-controlled clinical trial was to evaluate the effectiveness of the addition of raloxifene to GnRH analogues on uterine, leiomyoma, and non-leiomyoma sizes, and on the occurrence of leiomyoma-related symptoms. METHODS: After randomization using a computer-generated list, 100 pre-menopausal women with symptomatic uterine leiomyomas received either leuprolide acetate depot plus raloxifene 60 mg daily (group A) or leuprolide plus placebo tablet (group B) for six cycles of 28 days. At baseline and after treatment, uterine, leiomyoma and non-leiomyoma sizes, and leiomyoma-related symptoms were evaluated for each woman. Analysis was by intention-to-treat method. RESULTS: After six cycles of treatment, a significant decrease in uterine, leiomyoma, and non-leiomyoma sizes was detected in both groups in comparison with baseline. At the same time, no significant difference in uterine and non-leiomyoma sizes was observed between the groups. Leiomyoma sizes were significantly (P < 0.05) lower in group A than in group B. No difference was observed in leiomyoma-related symptoms between groups throughout the study period. CONCLUSIONS: In women treated with GnRH analogue, the raloxifene administration induces a higher reduction of leiomyoma sizes.     

Coexpression of Aspartic Proteinases and Human Leukocyte Antigen-DR in Human Transplanted Lung

Aspartic proteinases have recently been shown to be implicated in antigen processing. We explored the expression of two aspartic proteinases, cathepsins E and D, and of human leukocyte antigen-DR (HLA-DR) molecules in a consecutive series of 80 transbronchial biopsies from transplanted lungs. For controls, we studied five normal donor lungs (not suitable for transplantation on account of thoracic trauma) and macroscopically normal areas of three cancer-affected lungs. Two of the five unsuitable donor lungs showed minimal inflammatory changes. Macroscopically normal samples from the three cancerous lungs showed mild and focal inflammatory infiltrates. In histologically normal lungs, HLA-DR expression was limited to professional antigenpresenting cells. Macroscopically normal lung samples with minimal inflammatory changes from both donor and cancer lungs showed variable HLA-DR expression by alveolar and bronchial epithelial cells and by endothelial cells. All transplanted lung biopsies showed HLA-DR expression by epithelial (alveolar and bronchial) and endothelial cells, with a trend for increased positivity in acute rejection. Cathepsin E was restricted to Clara and to rare bronchus-associated lymphoid tissue-related epithelial cells in histologically normal lung samples, whereas minimal de novo cathepsin E expression by rare alveolar pneumocytes was noted in control lung samples exhibiting minimal inflammatory changes. In all transplanted lung biopsies, cathepsin E was diffusely expressed de novo by hyperplastic alveolar epithelial cells, regardless of the presence or degree of rejection. Cathepsin D was expressed only by alveolar macrophages and by ciliated bronchial cells of normal, minimally inflamed, and transplanted lungs. In transplanted lung, Clara cells and several hyperplastic alveolar pneumocytes coexpressed HLA-DR and cathepsin E, whereas all alveolar macrophages and a few ciliated cells coexpressed cathepsin D and HLA-DR The present investigation suggests that the de novo expression of cathepsin E and HLA-DR by hyperplastic alveolar pneumocytes of transplanted lung may be crucial for antigen processing and presentation to recipient competent T cells, and thus for the triggering of the immune-inflammatory cascade that leads to rejection.     

Neural control of fast-regular saccades and antisaccades: an investigation using positron emission tomography

 Regional cerebral blood flow changes related to the performance of two oculomotor tasks and a central fixation task were compared in ten healthy human subjects. The tasks were: (a) performance of fast-regular saccades; (b) performance of voluntary antisaccades away from a peripheral cue; (c) passive maintenance of central visual fixation in the presence of irrelevant peripheral stimulation. The saccadic task was associated with a relative increase in activity in a number of occipitotemporal areas. Compared with both the fixation and the saccadic task, the performance of antisaccades activated a set of areas including: the superior and inferior parietal lobules, the precentral and prefrontal cortex, the cingulate cortex, and the supplementary motor area. The results of the present study suggest that: (a) compared with self-determined saccadic responses the performance of fast regular, reflexive saccades produces a limited activation of the frontal eye fields; (b) in the antisaccadic task the inferior parietal lobes subserve operations of sensory-motor integration dealing with attentional disengagement from the initial peripheral cue (appearing at an invalid spatial location) and with the recomputation of the antisaccadic vector on the basis of the wrong (e.g., spatially opposite) information provided by the same cue. Received: 20 May 1996 / Accepted: 28 January 1997