The histone deacetylase inhibitor ITF2357 has anti-leukemic activity in vitro and in vivo and inhibits IL-6 and VEGF production by stromal cells.

We have investigated the activity of ITF2357, a novel hydroxamate histone deacetylase inhibitor, on multiple myeloma (MM) and acute myelogenous leukemia (AML) cells in vitro and in vivo. ITF2357 induced apoptosis in 8/9 MM and 6/7 AML cell lines, as well as 4/4 MM and 18/20 AML freshly isolated cases, with a mean IC(50) of 0.2 microM. ITF2357 activated the intrinsic apoptotic pathway, upregulated p21 and downmodulated Bcl-2 and Mcl-1. The drug induced hyperacetylation of histone H3, H4 and tubulin. When studied in more physiological conditions, ITF2357 was still strongly cytotoxic for the interleukin-6 (IL-6)-dependent MM cell line CMA-03, or for AML samples maximally stimulated by co-culture on mesenchymal stromal cells (MSCs), but not for the MSCs themselves. Interestingly, ITF2357 inhibited the production of IL-6, vascular endothelial growth factor (VEGF) and interferon-gamma by MSCs by 80-95%. Finally, the drug significantly prolonged survival of severe combined immunodeficient mice inoculated with the AML-PS in vivo passaged cell line already at the 10 mg/kg oral dose. These data demonstrate that ITF2357 has potent anti-neoplastic activity in vitro and in vivo through direct induction of leukemic cell apoptosis. Furthermore, the drug inhibits production of growth and angiogenic factors by bone marrow stromal cells, in particular IL-6 and VEGF.

Detection of Babesia caballi in Amblyomma variegatum ticks (Acari: Ixodidae) collected from cattle in the Republic of Guinea.

A reverse line blot hybridisation (RLB) assay was applied to screen Amblyomma variegatum adult ticks (n = 504) collected from N'Dama cattle in the Republic of Guinea. In a PCR, the V1 hypervariable region of the 16S ribosomal RNA (rRNA) gene was amplified with a set of primers unique for species of the genera Anaplasma and Ehrlichia, and the V4 hypervariable region of the 18S rRNA gene was amplified with primers specific for members of the genera Theileria and Babesia. Amplified PCR products from A. variegatum ticks were hybridised onto a membrane, to which oligonucleotide probes species-specific for Ehrlichia/Anaplasma and Theileria/Babesia parasites were covalently linked. No pathogens belonging to Ehrlichia/Anaplasma species were found, while 10 DNA samples resulted positive for Babesia caballi and 5 samples for Theileria velifera. This is the first report of B. caballi in A. variegatum ticks. One of the B. caballi positive samples was sequenced. This new strain (BcabGuinea) showed a 97% similarity to the Z15104 B. caballi GenBank sequence.

Trichinella britovi etiological agent of sylvatic trichinellosis in the Republic of Guinea (West Africa) and a re-evaluation of geographical distribution for encapsulated species in Africa.

In West Africa, Trichinella infection was documented in humans and animals from Senegal in the 1960s, and the biological characters of one isolate showed a lower infectivity to domestic pigs and rodents when compared with that of a Trichinella spiralis pig isolate from Europe. To identify the Trichinella species present in West Africa, a survey was conducted in a total of 160 wild animals in the Republic of Guinea. Three Viverridae, one true civet (Viverra civetta) and two African palm civets (Nandinia binotata) from the Fouta Djallon Massif, Pilimini Subprefecture, were found positive by artificial digestion of muscle samples. Trichinella larvae from these three viverrids were identified as Trichinella britovi and no difference was detected in three examined sequences from these African isolates and the reference strain of T. britovi from Europe, indicating common ancestry, an historically continuous geographic distribution, and recent isolation for African and European populations. The detection of T. britovi in West Africa modifies our knowledge about the distribution of encapsulated species of Trichinella in Africa. Thus, Trichinella nelsoni is now considered to have a distribution limited to the Eastern part of the Afrotropical region from Kenya to South Africa. This provides a plausible explanation for the presence of Trichinella T8 in Namibia and South Africa, and further suggests that T. britovi could be the Trichinella species circulating among wild animals of Northern Africa.

HIV/HCV co-infection: natural history.

HIV and HCV share common transmission pathways, but HCV is more efficiently transmitted through blood than with sexual exposure. Thus HCV coinfection is frequent in HIV seropositives, mainly in those with history of injection drug use and/or transfusion. HIV coinfection increases HCV replication rate, the rate of HCV vertical transmission and accelerates the course of hepatitis C towards cirrhosis and hepatocellular carcinoma. The evidence of an effect of HCV on HIV disease progression is less convincing. The results of several studies suggest that HCV coinfection does not hasten the progression of HIV infection towards AIDS. However two recent studies showed that HCV coinfection is independently associated with a lower restoration of CD4 counts during combination antiretroviral treatment. However this finding should be confirmed by additional studies.

In vivo exposure to NO2 reduces TNF and IL-6 production by endotoxin-stimulated alveolar macrophages.

Exposure to NO2 appears to affect lung defense mechanisms. We exposed rats to 10 ppm of NO2 for 24 h or 7 days and studied the production of tumor necrosis factor (TNF), interleukin-6 (IL-6), and prostaglandin E2 (PGE2) by alveolar macrophages after endotoxin stimulation. TNF and IL-6 production was significantly decreased (four-to sixfold) in the cell lysate of alveolar macrophages isolated from rats exposed to NO2. In parallel, PGE2 production was significantly increased in the same samples and in the bronchoalveolar lavage fluid. Northern blot analysis of the two cytokines indicated a reduction of the mRNA content. We also studied the expression of the TNF receptor type 1 (TNF-R1), known to neutralize TNF activity in its soluble form, and found that expression of the mRNA was increased after endotoxin stimulation. We can conclude that rats exposed to NO2 produce less TNF and IL-6 and that this might be related to increased PGE2 production and increased expression of TNF-R1.

Biochemical effects of acute and subacute nitrogen dioxide exposure in rat lung and bronchoalveolar lavage fluid.

The pulmonary inflammatory response to NO2 exposure was measured by evaluating a series of biochemical and cellular parameters in rat bronchoalveolar lavage fluid. Animals were exposed to 9 mg/m3 (5 ppm) or 18 mg/m3 (10 ppm) of the gas for 24 h or 7 days. After bronchoalveolar lavage collection, a differential count of polymorphonuclear leukocytes, macrophages, and lymphocytes was done. A significant increase in polymorphonuclear leukocytes was found after 24 h of exposure, and after 7 days the number of macrophages increased significantly. After 7 days of exposure to 9 mg/m3 of NO2 (a dose that under our conditions did not induce migration of cells in the bronchoalveolar spaces) the ex vivo phorbol myristate acetate-induced superoxide anion production by resident cells was inhibited. After 24 h and 7 days of exposure to 18 mg/m3 of NO2, phorbol myristate acetate-induced superoxide anion production was lower than in the control group. The migration of polymorphonuclear leukocytes in the bronchoalveolar lavage fluid was not associated with any real increase in elastase. However, there was a dose- and time-dependent increase in alpha 1-proteinase inhibitor in response to both 9 and 18 mg/m3 of NO2. Total glutathione was significantly increased in blood by 24 h treatment with 9 or 18 mg/m3 of NO2, whereas blood oxidized glutathione was not affected. In lung tissue we observed only a significant increase of oxidized glutathione after 24 h of exposure to 9 and 18 mg/m3 of NO2. These data suggest that many biochemical and cellular parameters are altered after acute or subacute exposure to relatively high doses of NO2, especially in the first 24 h.(ABSTRACT TRUNCATED AT 250 WORDS)

An automated system for animal exposure to nitrogen dioxide.

We have designed an automated system for controlled exposure of rodents to nitrogen dioxide (NO2). The system consists of 1) two stainless-steel exposure chambers (3.96 m3), for control and treated animals, in which temperature, humidity, and air changes are consistent with current guidelines for rodent housing; 2) a cylinder containing 1% NO2 in N2; 3) a detector connected with a personal computer system that monitors the NO2 concentration, and regulates the NO2 influx, using a closed-loop feedback; and 4) a fan system to distribute the gas uniformly within the chambers. In a typical experiment rats were exposed to 15 ppm of NO2, and changes in the differential count of polymorphonuclear leukocytes, macrophages, and lymphocytes in bronchoalveolar lavage fluid were investigated.

O6-alkylguanine DNA alkyltransferase is induced by human recombinant interferon-alpha A/D in mouse liver.

Treatment of C57Bl or BALB/C mice with human interferon-alpha A/D (HuIFN-alpha A/D) significantly increased hepatic levels of the DNA repair enzyme O6-alkylguanine DNA alkyltransferase (AT). The maximum induction was seen 24 h after a single dose of 50-100 micrograms/kg IFN-alpha A/D. No induction was observed in rat liver hepatocytes cultured in vitro. Liver AT was also induced by poly(I:C), which is a potent IFN inducer. By increasing AT levels, IFN could protect against the potentially mutagenic alkylation at guanine O6 position caused by some carcinogens. Moreover this finding suggests a link between immune response and the DNA repair system, possibly acting in concert to defend the body from potentially toxic compounds.

O6-Alkylguanine-DNA alkyltransferase content in synchronised human cancer cells.

The DNA repair enzyme O6-alkylguanine-DNA alkyltransferase (AT) was analysed in the human ovarian-cancer SW626 cell line and in the human promonocytic leukemia U937 cell line following their synchronisation with low non-toxic concentrations of methotrexate. In SW626, AT increased in the early S phase of the cell cycle and then declined during progression of the S phase to levels found in the G1 phase of unsynchronised cells. In contrast, at the G1/S-phase boundary and in the S phase, U937 cells showed a lower AT content than did exponentially growing unsynchronised cells. In addition, AT activity was greatly reduced in resting U937 cells but was not reduced appreciably in resting SW937 cells but was not reduced appreciably in resting SW626 cells. The results of these studies indicate that AT fluctuations do not follow a constant pattern during the cell cycle of different cell lines.

Interferon inducers increase O6-alkylguanine-DNA alkyltransferase in the rat liver.

We investigated whether treatment with the interferon inducer polyinosinic-polycytidylic acid and other cytokines (interleukin-1, tumor necrosis factor) or the cytokine inducer lipopolysaccharide modified O6-alkylguanine-DNA alkyltransferase (AT) in rat liver. AT levels were determined in liver extracts using N-[3H]methyl-N-nitrosourea alkylated calf thymus DNA as substrate and an HPLC procedure to measure O6-methylguanine. Doses as low as 0.1 mg/kg i.p. of polyinosinic-polycytidylic acid caused a highly significant increase (P less than 0.01) in AT levels in the liver, evident either 24 or 48 h after treatment. Lipopolysaccharide at the dose of 80 micrograms/kg i.p. also induced AT whereas interleukin-1 (60 micrograms/kg) or tumor necrosis factor (60 micrograms/kg) were inactive. Treatment with human recombinant interferon alpha A/D caused a highly significant increase in AT levels, thus confirming the hypothesis that interferon was probably responsible for the observed effect. These results suggest a link between the immune response and DNA repair mechanisms.

Assessment of VVI diagnostic pacing mode in patients with cardioinhibitory carotid sinus syndrome.

In this study, we used Holter pacemakers in a group of 13 patients affected by severe carotid sinus syndrome in order to evaluate its evolution. All the patients had one to three syncopal episodes and frequent other symptoms such as fainting, dizziness, lightheadedness and pre-syncope interferring with their daily activity so that pacemaker therapy was considered necessary. Patient selection criteria were: presence of the isolated cardioinhibitory type, absence of associated sinus dysfunction and absence of symptomatic VVI pacemaker effect. All the patients received a Micropacer 1 device; among special functions, bradycardia events counter was activated and programmed so that each sequence of three consecutive beats at a cycle length 1.5 sec (i.e., 4.5 sec total interval) could be recognized and stored in its memory. The follow-up lasted 13 +/- 7 months. Brady events occurred in eight out of 13 patients (62%), during this period. Syncope and major symptoms disappeared in all the patients; mild dizziness recurred rarely in two patients and were not linked to brady-events recording. In conclusion, disappearance of severe symptoms observed after pacemaker implant in cardioinhibitory carotid sinus syndrome seems to depend from pacing therapy, in most cases, yet from the benign natural course of the disease in some other cases.

Multiple-drug chemotherapy for the primary treatment of osteosarcoma of the extremities.

Fifty-five cases of osteosarcoma of the extremities were treated between 1972 and 1976 by combined surgery and chemotherapy (vincristine, adriamycin and methotrexate in medium doses) for 18 months. The follow-up ranges from 30 to 80 months (mean = 48 months). Twenty-six patients remained free from any evidence of disease, two had local recurrences but no metastases and 27 had metastases (four of these also had local recurrences). In 12 patients, the metastases appeared after the end of chemotherapy. Both metastases and local recurrences were more frequent in patients who had segmental bone resection (7/8) than in those treated by more radical surgery (22/47). Comparison with an "historical" group (94 osteosarcoma patients treated by operation alone in our Institute between 1960 and 1971) showed that the percentage of patients free from evidence of disease was higher in the group who receiving chemotherapy. In addition, the appearance of metastases in this group was delayed (mean = 16 months) as compared with the historical controls (mean = 8 months). On the other hand, after the same kind of operative treatment, the rate of local recurrences and the time of their appearance was almost identical in both groups.

Adjuvant multiple drug chemotherapy for osteosarcoma of the extremity: a 6 year report.

Fifty-five cases of osteosarcoma of the extremities were treated between 1972 and 1976 with combination surgery and polychemotherapy (vincristine, adriamycin and methotrexate at medium doses) for 18 months. Their follow-up presently ranges between 30 and 80 months (mean = 48 months). Twenty-six patients remained free from disease signs, 2 showed local recurrence but no metastases, and 27 exhibited metastases (4 of these also had local recurrences). In 12 patients, the metastases appeared after the end of chemotherapy. Both metastases and local recurrences were more frequent in those patients submitted to segmental bone resection (7/8) than in those treated by more radical surgery (22/47). Comparison with a historical group (94 osteosarcoma patients treated with surgery alone at our Institute between 1960 and 1971) revealed that, during the follow-up period considered, the percentage of patients free from disease signs was higher in the group that also received chemotherapy. In addition, in this group metastatic appearance was delayed (mean = 15 months) as compared to historical controls (mean = 8 months). On the other hand, after the same kind of surgery, the rate of local recurrences and the time of their appearance was practically the same in both groups.