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Biomedical subjects

M Marani

Publications and source records attributed to M Marani.

At least 19 recordsLinked to original sources
PubMed2005

Human immunodeficiency virus infection: personal experience in changes in head and neck manifestations due to recent antiretroviral therapies.

Both the incidence and prevalence of human immunodeficiency virus infection are increasing in the world. Diseases of ENT districts are more frequent in human immunodeficiency virus-infected patients and involve all the otolaryngological sites. The otorhinolaryngological manifestations in association with HIV infection are mainly atypical, so common in the clinical practice, really aspecific and very frequent in ENT daily routine (such as sinusitis, otitis, etc.) and, therefore, immunodeficiency may not be suspected. In other cases, ENT evidence is more peculiar or unusual, such as opportunistic infections, rare neoplasm and tumours with an unusual course, giving a very high suspect of a human immunodeficiency virus-related infection. The most frequent malignant neoplasm is Kaposi's Sarcoma which is extremely rare in non-human immunodeficiency virus-infected subjects; the second most frequent is non-Hodgkin's lymphoma with 50% in extranodal sites (oral and maxillary sinus). Following a review of the literature, modifications caused by current antiretroviral treatment on head and neck manifestations of human immunodeficiency virus infection have been evaluated. Highly active antiretroviral therapy is a new therapeutic strategy, based on poly-chemo-therapeutic schemes, providing simultaneously two or more anti-retroviral drugs. We have used highly active antiretroviral therapy in human immunodeficiency virus infection since 1997, substituting previous mono-chemotherapy based on Zidovudine or Didanosine alone. Highly active antiretroviral therapy is extremely efficient in reducing the viral load of human immunodeficiency virus and increasing CD4+ T-lymphocyte count. These biological effects are associated with an improvement in immune functions. To evaluate the effects of highly active antiretroviral therapy on otorhinolaryngological manifestations in human immunodeficiency virus infection, we performed a retrospective study on 470 adults, observed over 14 years (1989-2002) and constantly receiving the same treatment, with follow-up from 7 to 80 months. A total of 250 subjects underwent mono-antiretroviral chemotherapy (1989-1996), while 220 underwent highly active antiretroviral therapy (1997-2002). The results of the retrospective study showed that highly active antiretroviral therapy has greatly improved the control of the immune-deficiency (increasing the range of CD4+), reducing the number of otorhinolaryngological manifestations (also tumours). On the other hand, 2 patients presented sudden unilateral hearing loss following treatment: toxicity due to association of new drugs cannot be excluded.

Adult↗
PubMed2003

Expression of Smac/DIABLO in ovarian carcinoma cells induces apoptosis via a caspase-9-mediated pathway.

We have constructed Ad CMV-Smac, a recombinant adenovirus encoding Smac/DIABLO, the recently described second mitochondrial activator of caspases. Transfection of ovarian carcinoma cells with Ad CMV-Smac at multiplicities of infection of 3-60 pfu/cell leads to increasing apoptosis in a dose-dependent manner. Western blot analysis confirms that Smac-induced apoptosis proceeds via a pathway mediated primarily by caspase-9 that can be inhibited by zLEHD-fmk and overexpression of the X-linked inhibitor of apoptosis protein (XIAP). In contrast, there is no cleavage of either caspase-8 or caspase-12. Ad CMV-Smac appears to induce apoptosis independently of cytochrome c release from mitochondria and is not inhibited by overexpression of Bcl-2. Ad CMV-Smac can combine with other proapoptotic factors, such as cisplatin, paclitaxel, and procaspase-3, to produce greater levels of apoptosis in transfected cells.

Antineoplastic Agents↗
PubMed2001

Overexpression of BCL-X(L) underlies the molecular basis for resistance to staurosporine-induced apoptosis in PC-3 cells.

We have reported previously that among human prostate cancer cell lines LNCaP but not PC-3 cells undergo apoptosis after treatment with the protein kinase inhibitor staurosporine (STS). We have now further investigated this model to uncover the molecular mechanism causing resistance to STS-induced apoptosis in PC-3 cells. S-100 lysates of both cell lines showed biochemical changes typical of apoptosis after the addition of cytochrome c and dATP, suggesting that the postmitochondrial phase of apoptosis was intact. Upon addition of STS, the proapoptotic molecules Bax and Bad became predominantly mitochondrial in both cell lines. This, in turn, was followed by loss of mitochondrial transmembrane potential, translocation of cytochrome c to the cytosol, activation of caspase-9, -3, and -7, and cleavage of the apoptotic targets, DNA fragmentation factor and poly(ADP-ribose) polymerase, in LNCaP but not in PC-3 cells. Components of the mitochondrial permeability transition pore, adenine nucleotide transporter and voltage-dependent anion channel, were normally expressed in the correct subcellular fraction of both cell lines. Overexpression of the proapoptotic proteins Bax and Bad, fused to a green fluorescent protein but not of green fluorescent protein alone, induced apoptosis in >80% of PC-3 cells. These experiments suggested that a factor protecting the mitochondria of PC-3 cells mediates resistance to STS-induced apoptosis. A wide search among the antiapoptotic Bcl-2 family members was performed, and Bcl-X(L) was found to be overexpressed in PC-3 cells. Experiments down-regulating Bcl-X(L) expression by using the tyrosine kinase inhibitor genistein, sodium butyrate, or an antisense Bcl-X(L) oligonucleotide restored sensitivity to apoptosis in PC-3 cells. Thus, Bcl-X(L) overexpression is one of the mediators of resistance to STS-induced apoptosis in the prostate cancer cell line PC-3.

Apoptosis↗
PubMed2001

Adenovirus-mediated Bax overexpression for the induction of therapeutic apoptosis in prostate cancer.

Using adenoviral technology, we overexpressed the proapoptotic molecules pro-caspase-3, pro-caspase-7, and Bax to induce therapeutic apoptosis of prostate cancer cell lines growing in vitro and in vivo. Because overexpressed pro-caspase-3 did not undergo autocatalytic activation in any of the five prostate cancer cell lines evaluated, this strategy was unable to engage any component of the apoptotic pathway. Overexpressed pro-caspase-7 was proteolytically cleaved in LNCaP and LnCaP-Bcl-2 cells but not in PC-3, DU-145, or TsuPr(1) cells. Cleavage was associated with engagement of many components of the apoptotic pathway, including DEVDase activity, cleavage of intracellular caspase targets such as the DNA fragmentation factor and the proapoptotic Bid, release of cytochrome c from the mitochondria to the cytoplasm, and terminal deoxynucleotidyl transferase-mediated nick end labeling. No apoptosis was observed in the cells where caspase-7 did not undergo autocatalytic activation. Searching for an approach that would more reliably induce therapeutic apoptosis of prostate cancer cell lines, we used a binary adenoviral system to overexpress the proapoptotic molecule Bax. Bax was dramatically overexpressed and caused apoptosis of every cell line infected by engaging the mitochondrial pathway, including proteolytic cleavage and catalytic activation of the caspases, cleavage of caspase substrates, release of cytochrome c from the mitochondria, and DNA fragmentation. Furthermore, three injections of the Bax overexpression system into PC-3 cell tumors in nude mice in vivo caused a 25% regression in tumor size corresponding to a 90% reduction relative to continued tumor growth in animals that received injections with the control binary system expressing Lac-Z. These experiments show that adenovirus-mediated Bax overexpression is capable of inducing therapeutic programmed cell death in vitro and in vivo by activating the mitochondrial pathway of apoptosis. On the basis of these studies, we conclude that manipulation of Bax expression is an attractive new gene therapy approach for the treatment of prostate cancer.

Adenoviridae↗
PubMed2001

Pro-caspase-3 overexpression sensitises ovarian cancer cells to proteasome inhibitors.

The ubiquitin-proteasome pathway plays a critical role in the degradation of several proteins involved in the cell cycle. Dysregulation of this pathway leads to inhibition of cellular proliferation and the induction of apoptosis. Ubiquitination and its downstream consequences have been investigated intensively as targets for the development of drugs for tumour therapy. Here we have investigated the mechanism of apoptosis induced by the proteasome inhibitors MG-132, lactacystin and calpain inhibitor I (ALLN), in the HEK 293 cell line and the ovarian cancer cell lines SKOV3 and OVCAR3. We have found strong caspase-3-like and caspase-6-like activation upon treatment of HEK 293 cells with MG-132. Using a tricistronic expression vector based on a tetracycline-responsive system we generated stable SKOV3 nd OVCAR3 cell lines with inducible expression of pro-caspase-3. Induction of pro-caspase-3 expression in normally growing cells does not induce apoptosis. However, in the presence of the proteasome inhibitors MG-132, lactacystin or ALLN we found that cells overexpressing pro-caspase-3 are rapidly targeted for apoptosis. Our results demonstrate that pro-caspase-3 can sensitise ovarian cancer cells to proteasome inhibitor-induced apoptosis, and a combination of these approaches might be exploited for therapy of ovarian and other cancers.

Acetylcysteine↗
PubMed2001

Herpes simplex virus thymidine kinase/ganciclovir-induced cell death is enhanced by co-expression of caspase-3 in ovarian carcinoma cells.

There is a need to enhance the efficacy of genetic prodrug activation therapy using herpes simplex virus thymidine kinase (tk) and ganciclovir (GCV) following disappointing results in early clinical trials. tk/GCV has been shown to lead to the activation of caspase-3, a potent executor of apoptosis. We demonstrate that co-expression of pro-caspase-3 with tk/GCV leads to enhanced cell death in ovarian carcinoma cells in vitro. Following transfection with recombinant adenoviral vectors encoding tk, GCV treatment leads to greater cell death in pro-caspase-3-expressing clones of SKOV3 and IGROV1 than control cells, as well as more rapid activation of caspase-3 and more rapid cleavage of PARP. Flow cytometry suggests that there is a greater degree of S-phase block in the pro-caspase-3-expressing clones than in control cells following treatment with tk/GCV. None of these effects is seen following transfection with a control adenovirus that does not encode tk. The increased cell death, early caspase-3 activation and PARP cleavage, and flow cytometric changes seen in pro-caspase-3-expressing cells can be partially inhibited by treatment with benzyloxycarbonyl-val-ala-asp fluoromethylketone, a synthetic caspase inhibitor. Our data suggest that co-expression of pro-caspase-3 may lead to a significant enhancement of the efficacy of tk/GCV therapy.

Adenoviridae↗
PubMed2000

Heterogeneous apoptotic responses of prostate cancer cell lines identify an association between sensitivity to staurosporine-induced apoptosis, expression of Bcl-2 family members, and caspase activation.

BACKGROUND: The goal of this work was to identify mechanisms for the inability of metastatic prostate cancer cells to engage the apoptotic pathway following hormonal or cytotoxic therapy. METHODS: Genotypically diverse cell lines isolated from patients with metastatic disease were used. RESULTS: The LNCaP and TsuPr(1) lines exhibited quintessential apoptotic features in response to the pleiotropic apoptotic inducer staurosporine (STS): rapid cytochrome c translocation to the cytosol, proteolytic processing and catalytic activation of caspase-3 and -7, proteolytic inactivation of the death substrates DNA fragmentation factor (DFF) and poly-ADP-ribose polymerase (PARP), and TUNEL-positive polyfragmented nuclei. In contrast, DU-145 and PC-3 cells exhibited few, if any, of these features, while appearing necrotic by confocal microscopy. The presence of caspase-3 and -7 without proteolytic processing suggested that the apoptotic blockade was upstream of executioner caspases in these resistant cell lines. To identify the locus of this block, Western blot analysis of cytochrome c subcellular localization and of pro- and antiapoptotic Bcl-2 family members was performed, and suggested that heterogeneous expression of these proteins might be the underlying mechanism for apoptotic resistance to STS in these cell lines. Thus, the absence of the proapoptotic Bax in DU-145 cells indicated a mechanism for apoptotic resistance of these cells. Similarly, decreased Bax expression during STS treatment, coupled with overexpression of the antiapoptotic Bcl-x(L) and inability to translocate cytochrome c to the cytosol, provided a mechanism for the insensitivity of PC-3 cells. CONCLUSIONS: These observations suggest that activation of the apoptotic machinery in metastatic prostate cancer cell lines may be determined by expression levels of Bcl-2 family members, by the ability of cytochrome c to translocate to the cytosol, and by the ability of the caspase pathway to react in response to activation of the mitochondrial phase.

Apoptosis↗
PubMed2000

Validation of the 13C-urea breath test for the diagnosis of Helicobacter pylori infection in children: a multicenter study.

OBJECTIVE: The 13C-urea breath test (13C-UBT) is a safe, noninvasive, and accurate test for the detection of Helicobacter pylori (H. pylori) infection in adults. The aim of this study was to evaluate sensitivity and specificity of 13C-UBT in children using different types of test meal, doses of 13C-urea and breath sampling intervals. As yet, a validated, standardized 13C-UBT protocol for children has not been formulated. METHODS: 13C-UBT was performed in 115 children and repeated within 3 days, modifying the test meal or the dose of 13C-urea. H. pylori status was assessed by histology and rapid urease test. 13C-UBT was performed using 100 mg or 50 mg of 13C-urea and a fatty test meal (100 FA; 50 FA), 50 mg of 13C-urea, and a carbohydrate test meal (50 CA). Breath samples were collected every 10 min for 60 min. RESULTS: The 13C-UBT in children was highly sensitive and specific with all three protocols used. The best combination of sensitivity (97.92%) and specificity (97.96%) was obtained with Protocol 50 FA at 30 min with a cut-off of 3.5 per mil. CONCLUSIONS: The 13C-UBT is an accurate test for the detection of H. pylori infection also in children. Administration of 50 mg of 13C-urea, a fatty test meal, and breath sampling at 30 min appears to be the most convenient protocol.

Adolescent↗
PubMed2000

Induction of apoptosis in BPH stromal cells by adenoviral-mediated overexpression of caspase-7.

PURPOSE: We hypothesized that expression/activity of critical components of the apoptotic pathway can be used to induce apoptosis of a human prostate cell line derived from benign prostatic hyperplasia (BPH) tissue. MATERIALS AND METHODS: We analyzed the apoptotic pathway in BPH cells treated with the powerful inducer of apoptosis, staurosporine (STS), and adenoviruses overexpressing caspase-3, -7, or the control gene lacZ. RESULTS: Twelve hours post-STS, most BPH cells were floating in the culture medium, TUNEL staining was widespread, and DEVDase activity (the catalytic activity of type II caspases) was increased. The pan-caspase inhibitor, Z-VAD-FMK, prevented STS-induced apoptosis. Based on these observations, we performed immunoblot analysis for the three known group II caspases (that is caspase-2, -3 and -7), but none of them was detected with three commercially available antibodies. Nevertheless, in view of the presence of increased DEVDase activity, we reasoned that a group II caspase must be a critical mediator of apoptosis in this model. If correct, we postulated that overexpression and activation of a type II caspase should cause apoptosis. To test this hypothesis, we coupled the cDNAs encoding caspase-3 and caspase-7 to adenoviral vectors and obtained constructs AvC3 and AvC7. Cells infected with AvC3 or AvC7 overexpressed the protein for caspase-3 or -7 within 24 to 48 hours. Caspase-3 overexpression did not cause apoptosis above that observed in cells receiving the control adenovirus expressing the lacZ cDNA (AvLac-Z). In contrast, caspase-7 overexpression induced massive apoptosis. BPH cells were then infected with increasing multiplicity of infection (MOI) of AvC7 and AvlacZ. A positive correlation was found between the amount of caspase-7 expressed and the level of DEVDase activity measured. AvC7 at MOIs of 25:1 and 50:1 induced apoptosis in about 50% of BPH cells at 72 hours post-infection. This effect was AvC7 specific, because the same MOIs of AvlacZ were not apoptogenic. CONCLUSIONS: Adenoviral-mediated overexpression of caspase-7 induces apoptosis of BPH-derived cells.

Adenoviridae↗
PubMed2000

[Predictive factors of etiology (bacterial or viral) in acute gastroenteritis in childhood].

The aim of the study was to identify immediate clinical and/or laboratory findings able to differentiate bacterial from viral etiology of acute gastroenteritis in pediatric patients. We studied 52 children, aged between 5 months and 12 years, consecutively admitted to hospital with acute diarrhoea lasting less than 5 days. All the patients were divided into 4 groups according to etiologic agent, subsequently demonstrated by culture: salmonellae (group A), rotavirus (group B); combined salmonellae and rotavirus (group C) and no pathogen (group D). The contemporary presence of fever > 39 degrees C, number of daily liquid stools > 6, presence of bloody diarrhoea, positivity of C-reactive protein and hyponatremia (< 135 mEq/l) allowed to recognize the etiology (viral or bacterial) before results of culture (sensitivity was 71% and specificity was 97%). In particular, hyponatremia resulted significantly lower in group A and C than in group B and D. We concluded that hyponatremia can be considered a marker for acute gastroenteritis caused by salmonellae.

Acute Disease↗
PubMed1999

Failures in snoring and OSAS surgery.

Significant reduction of snoring noise and valid prevention of neurological and/or cardiovascular complications of OSAS are the basic goals of all modern snoring and OSAS surgical procedures. Any kind of operation, single or multiple, included into a one-step or multistep programs, is said to fail if snoring is not reduced to a significant extent for the patient or if clinical and/or instrumental data after the operation show that Upper Airways Resistance Syndrome (UARS) or OSAS continues to be probably dangerous for the patient to some extent. The real figure of failures in different situations of sleep-disordered syndromes surgery is discussed, along with the possible explanations. A group of patients operated on for snoring and OSAS in our clinic is analyzed retrospectively from the subjective point of view and by means of sleep studies to get a precise quantitative and qualitative idea of the failed cases. The final goal would be to understand how it is possible to reduce to a minimal level the number of true failures.

Adult↗
PubMed1996

The coeliac iceberg in Italy. A multicentre antigliadin antibodies screening for coeliac disease in school-age subjects.

BACKGROUND: Recent studies suggest that coeliac disease (CD) is one of the commonest, life-long disorders in Italy. The aims of this multicentre work were: (a) to establish the prevalence of CD on a nationwide basis; and (b) to characterize the CD clinical spectrum in Italy. PATIENTS AND METHODS: Fifteen centres screened 17,201 students aged 6-15 years (68.6% of the eligible population) by the combined determination of serum IgG- and IgA-antigliadin antibody (AGA) test; 1289 (7.5%) were IgG and/or IgA-AGA positive and were recalled for the second-level investigation; 111 of them met the criteria for the intestinal biopsy: IgA-AGA positivity and/or AEA positivity or IgG-AGA positivity plus serum IgA deficiency. RESULTS: Intestinal biopsy was performed on 98 of the 111 subjects. CD was diagnosed in 82 subjects (75 biopsy proven, 7 not biopsied but with associated AGA and AEA positivity). Most of the screening-detected coeliac patients showed low-grade intensity illness often associated with decreased psychophysical well-being. There were two AEA negative cases with associated CD and IgA deficiency. The prevalence of undiagnosed CD was 4.77 x 1000 (95% CI 3.79-5.91), 1 in 210 subjects. The overall prevalence of CD, including known CD cases, was 5.44 x 1000 (95% CI 4.57-6.44), 1 in 184 subjects. The ratio of known to undiagnosed CD cases was 1 in 7. CONCLUSIONS: These findings confirm that, in Italy, CD is one of the most common chronic disorders showing a wide and heterogeneous clinical spectrum. Most CD cases remain undiagnosed unless actively searched.

Adolescent↗
PubMed1992

Precision of gestational age assessment in the neonate.

The precision of the Ballard scale for assessing gestational age (GA) was evaluated in a consecutive sample of 227 preterm and/or low-birth-weight neonates. Each newborn was rated independently by two neonatologists and the difference in GA estimation between them was computed. The estimated precision was not high, the 95% tolerance interval estimate being as large as 7.4 weeks. The precision of the neurologic and physical parts of the scale was poorer than that of the complete scale (95% of differences less than 10.5 and 9.2 weeks respectively), and more influenced by the type of delivery. These findings are not unexpected from statistical theory, and cast doubts on the use of only the physical part of the Ballard scale in assessing GA, since greater accuracy could be accompanied by reduced precision.

Anthropometry↗
PubMed1981

Ph1-negative T lymphocytic colonies in agar cultures of peripheral blood in chronic myeloid leukemia.

T lymphocytic colony formation by peripheral lymphocytes separated by discontinuous albumin gradient centrifugation was evaluated in 8 patients with Philadelphia (Ph1)-positive chronic myeloid leukemia (CML). Colonies were obtained using a liquid-on-agar culture system recently introduced (PHA overlayer-leukocyte feeder layer assay) which has been shown to be simple and reliable. The pattern of colony growth in CML and in normal controls was similar, the peak ranging from the 4th to the 6th day. Also the morphological aspects of colonies did not differ in the two groups. Cells recovered from CML lymphocytic colonies were shown to belong to T cell lineage, as they are able to form spontaneous E-rosettes and to respond to mitogenic stimulation in vitro. In contrast, cells recovered from all other cultured fractions failed to display these properties. Cytogenetic analysis showed that T colony cells were Ph1-negative whereas the chromosome anomaly was found in nonlymphoid colonies of the same patients, thus suggesting a nonclonal origin of T lymphocytes in CML.

Agar↗
PubMed1981

[Hypertrophic exudative gastropathy in a child. A case report].

Case report of a four-year-old atopic boy with protein-losing hypertrophic gastropathy. The diagnosis was confirmed radiologically and with radioisotope examinations. Already after one week of a high protein diet, spontaneous remission occurred, accompanied by an asthmatic attack.

Child, Preschool↗
PubMed1980

[Growth in vitro of human bone marrow after freezing in liquid nitrogen: analysis of a case].

Human marrow is frozen according to the Schafer's technique; CFU-C proliferation and differentiation is then evaluated in relation to the time of storage in liquid nitrogen and to the time of in vitro culture in agar system according to the Pike and Robinson's technique. The highest plating efficiency is observed at the twelfth day of culture while in fresh cell suspension the growth peaks at the eighth day.

Bone Marrow Cells↗