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1.
J Med Microbiol ; 49(4): 339-342, 2000 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-10755627

RESUMO

Mycobacterium leprae multiplies within host macrophages. The mechanism of internalisation of the bacteria by the phagocytic cells is unknown. In this study, M. leprae was purified from the foot pads of experimentally infected nu/nu mice. Peritoneal macrophages were harvested from BALB/c mice or C57 beige (bg/bg) mice. The effect of protein kinase inhibitors (erbstatin, genistein or staurosporine for BALB/c and bg/bg mice, plus herbimycin for bg/bg mice) on phagocytosis of the mycobacteria by the macrophage monolayers was tested. The untreated (control) macrophages phagocytosed M. leprae. Phagocytosis by BALB/c macrophages was inhibited by erbstatin and staurosporine but not by genistein; all the protein kinase inhibitors prevented uptake of M. leprae by bg/bg cells. The results demonstrate that protein kinase regulates phagocytosis of M. leprae by macrophages. The mechanism might prove to be a rational drug target for mycobacteria that multiply intracellularly.


Assuntos
Macrófagos Peritoneais/imunologia , Mycobacterium leprae/imunologia , Fagocitose/fisiologia , Proteínas Quinases/fisiologia , Animais , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Genisteína/farmacologia , Hidroquinonas/farmacologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fagocitose/efeitos dos fármacos , Inibidores de Proteínas Quinases , Estaurosporina/farmacologia
2.
Int J Antimicrob Agents ; 13(2): 133-5, 1999 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-10595573

RESUMO

The resistance of mycobacteria to beta-lactam antibiotics is attributed to their ability to synthesize beta-lactamase. In our previous studies, beta-lactam/beta-lactamase-inhibitor combinations suppressed the growth of several mycobacteria in axenic cultures and ampicillin/sulbactam was bactericidal to Mycobacterium tuberculosis H37Rv in vitro, and to Mycobacterium leprae multiplying in mouse foot-pads. Since both these organisms multiply in phagocytic cells in the host, it is important to know whether the drug combination is active against mycobacteria multiplying in macrophages. We tested the action of ampicillin/sulbactam against four potentially pathogenic (to humans or to animals) mycobacteria, M. simiae, M. haemophilum, M. avium, M. microti, when phagocytosed by mouse macrophages. Bacteria were exposed to monolayers of peritoneal macrophages harvested from BALB/c mice. Unphagocytosed bacilli were removed and three concentrations of ampicillin/sulbactam were tested. Optimum activity was observed at 100 mg/l which killed 58-97% of the mycobacteria within macrophages, as determined by the CFU. beta-Lactam/beta-lactamase-inhibitors, especially ampicillin/sulbactam, might provide an effective alternative therapy against infections caused by mycobacteria resistant to other drugs.


Assuntos
Ampicilina/farmacologia , Inibidores Enzimáticos/farmacologia , Mycobacterium/efeitos dos fármacos , Penicilinas/farmacologia , Sulbactam/farmacologia , Inibidores de beta-Lactamases , Animais , Contagem de Colônia Microbiana , Interações Medicamentosas , Humanos , Técnicas In Vitro , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium/crescimento & desenvolvimento , Fagocitose
3.
J Antimicrob Chemother ; 44(2): 279-81, 1999 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10473236

RESUMO

We reported previously that an injectable form of ampicillin/sulbactam, Unasyn, was bactericidal to Mycobacterium leprae multiplying in mouse foot pads. In this study, we examined the effect of an orally active form of ampicillin/sulbactam, Sultamicillin, on the growth of M. leprae in mice. Three concentrations of the drug, mixed with the feed, were administered from the start until the mice were killed at 6 months; 0.01% of the drug inhibited bacterial growth by 54%, 0.10% by 74% and 0.20% by 93%. To test whether oral ampicillin/sulbactam was bactericidal, 0.50% of the drug, mixed with the feed, was administered to experimentally infected mice for 3 months during the logarithmic phase of bacterial growth, and then discontinued; multiplication of the bacilli was monitored monthly for the next 8 months. The results showed that orally active ampicillin/sulbactam is bactericidal to M. leprae.


Assuntos
Quimioterapia Combinada/farmacologia , Mycobacterium leprae/efeitos dos fármacos , Administração Oral , Ampicilina/administração & dosagem , Ampicilina/farmacologia , Animais , Quimioterapia Combinada/administração & dosagem , Membro Posterior/microbiologia , Hanseníase/tratamento farmacológico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Modelos Biológicos , Mycobacterium leprae/crescimento & desenvolvimento , Sulbactam/administração & dosagem , Sulbactam/farmacologia
7.
J Basic Microbiol ; 36(5): 341-9, 1996.
Artigo em Inglês | MEDLINE | ID: mdl-8914265

RESUMO

Lysophospholipids are key intermediates in the metabolism of phospholipids. Cytoplasmic membranes of both eukaryotes and prokaryotes are made of phospholipid bilayers. Phospholipases are activated during phagocytosis. Lysophospholipids generated by phospholipase A2 or A1 degrade cell membranes and can cause cell lysis. An active lysophospholipase, that hydrolyzes lysophospholipids, was detected by the radioisotope technique in Mycobacterium leprae. About two-thirds of the enzyme was particulate and one-third cytoplasmic. Optimum activity was at 37 degrees C, and at pH 6.0. Temperatures above 70 degrees C completely inactivated the enzyme. The compound AACOCF3, a trifluromethylketone analog of arachiodonic acid, inhibited the activity; the inhibition appeared to be of the uncompetetive type. The K(m) of the enzyme was 2.5 x 10(-4)M, suggesting a fairly strong affinity for the substrate. Lysophospholipids have been shown to be microbicidal to invading organisms. Possession of lysophospholipase by M. leprae is apparently one of the methods by which the bacilli overcome the defense mechanisms of the host.


Assuntos
Lisofosfolipase/metabolismo , Mycobacterium leprae/enzimologia , Ácidos Araquidônicos/farmacologia , Inibidores Enzimáticos/farmacologia , Concentração de Íons de Hidrogênio , Cinética , Lisofosfolipase/antagonistas & inibidores , Lisofosfolipídeos/metabolismo , Temperatura
11.
Indian J Lepr ; 65(2): 153-6, 1993.
Artigo em Inglês | MEDLINE | ID: mdl-8345228
12.
Microbios ; 76(309): 251-61, 1993.
Artigo em Inglês | MEDLINE | ID: mdl-8302203

RESUMO

Mycobacterium tuberculosis and Mycobacterium leprae develop resistance against the drugs used to treat tuberculosis and leprosy, respectively. Now multidrug-resistant tuberculosis is spreading in many countries, especially with the emergence of AIDS. Multidrug treatment is being promoted at present to eradicate leprosy. Since M. leprae may also become multidrug-resistant, new approaches have to be adopted for controlling mycobacterial diseases. Mycobacteria usually synthesize beta-lactamase and are insensitive to beta-lactam antibiotics. M. tuberculosis contains a constitutive beta-lactamase; de-repression of beta-lactamase has been reported in M. leprae. Three different beta-lactam/beta-lactamase-inhibitor combinations (ampicillin/sulbactam, amoxicillin/clavulanate and piperacillin/tazobactam) were used to suppress the growth of several strains of mycobacteria (including M. tuberculosis H37Rv) in vitro. Ampicillin/sulbactam is a potent bactericidal agent against M. leprae multiplying in mouse foot pads. In the present work, ampicillin/sulbactam showed higher activity than the other drug combinations. The beta-lactam/beta-lactamase inhibitors are likely to be effective as rational therapeutic agents against mycobacterial infections.


Assuntos
Antibacterianos/farmacologia , Quimioterapia Combinada/farmacologia , Mycobacterium/efeitos dos fármacos , Inibidores de beta-Lactamases , Amoxicilina/farmacologia , Combinação Amoxicilina e Clavulanato de Potássio , Ampicilina/farmacologia , Animais , Ácidos Clavulânicos/farmacologia , Camundongos , Testes de Sensibilidade Microbiana , Mycobacterium/enzimologia , Mycobacterium/crescimento & desenvolvimento , Ácido Penicilânico/análogos & derivados , Ácido Penicilânico/farmacologia , Piperacilina/farmacologia , Combinação Piperacilina e Tazobactam , Sulbactam/farmacologia
13.
Microbios ; 70(283): 139-44, 1992.
Artigo em Inglês | MEDLINE | ID: mdl-1323745

RESUMO

Neurotropism is one of the unusual properties of Mycobacterium leprae. The organism contains glutamic acid decarboxylase that generates gamma-amino-butyric acid (GABA) which is an inhibitory neurotransmitter. The binding of GABA by M. leprae in vitro was studied by using 3H-GABA as substrate. The bacteria had high-affinity binding sites for the amino acid. The uptake was a specific saturable process with a Km of 66.7 pM, pH optimum of 7.3 and a temperature optimum of 37 degrees C. The binding did not seem to be time-dependent, being complete in about 5 min. None of the known antagonists and agonists of GABA uptake by neurons, showed any significant effect on M. leprae; the receptors in the bacteria are apparently of a non-neuronal type, and different from those reported in spermatozoa and Pseudomonas.


Assuntos
Mycobacterium leprae/metabolismo , Ácido gama-Aminobutírico/metabolismo , Animais , Tatus , Concentração de Íons de Hidrogênio , Cinética , Receptores de GABA-A/efeitos dos fármacos , Temperatura , Fatores de Tempo
14.
Microbios ; 72(291): 137-42, 1992.
Artigo em Inglês | MEDLINE | ID: mdl-1287401

RESUMO

The multiplication of Mycobacterium leprae in foot pads of experimentally-infected mice was suppressed by intramuscular administration of ampicillin combined with sulbactam or YTR-830H, two potent inhibitors of beta-lactamase in the bacteria. The antibiotic or the inhibitors by themselves were inactive. Ampicillin/sulbactam also inhibited the growth of drug-resistant M. leprae which grew in the presence of rifampin or dapsone. The finding provides a new approach to treat leprosy and to overcome drug resistance of the mycobacteria.


Assuntos
Ampicilina/farmacologia , Mycobacterium leprae/efeitos dos fármacos , Sulbactam/farmacologia , Animais , Resistência Microbiana a Medicamentos , Quimioterapia Combinada/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium leprae/crescimento & desenvolvimento
15.
Indian J Lepr ; 63(3-4): 410-7, 1991.
Artigo em Inglês | MEDLINE | ID: mdl-1666638

RESUMO

My first contact with Dr. Dharmendra was through correspondence. While working for Ph.D., I wrote to him that a section in his book "Notes on Leprosy" was ambiguous. Instead of ignoring the letter, he replied, agreeing to clarify it in the revised edition. I went to work at Carville at the invitation of Dr. Kirchheimer, who had seen my Ph.D. thesis. Dr. Dharmendra visited Carville to receive the Damien-Dutton award and stayed there for a few days. Carville is an isolated place with no public transportation. I used to take him for afternoon drives to the countryside around Carville. He published some of our papers in Leprosy in India and later in Indian Journal of Leprosy. He was very prompt in acknowledging receipt of manuscripts and suggesting any changes to be made. He also reprinted in the Journal several of our papers published elsewhere, and also a lecture I gave at a meeting of the Japanese Leprosy Association. During one of my visits to India, Dr. M. C. Vaidya had arranged a talk by me at the All India Institute of Medical Sciences, New Delhi. At the invitation of Dr. Dharmendra, I visited him in his home. We used to exchange new year cards and letters. He wrote to me about his eye infection and consequent loss of sight in one eye. He asked me to write an editorial for an issue of Indian Journal of Leprosy (January 1989). The last time I met him was during the International Leprosy Congress held in New Delhi.


Assuntos
Mycobacterium leprae/enzimologia , Ampicilina/farmacologia , Animais , Glutamato Descarboxilase/metabolismo , Humanos , Hanseníase/tratamento farmacológico , Monofenol Mono-Oxigenase/metabolismo , Mycobacterium leprae/crescimento & desenvolvimento , Ácido Penicilânico/farmacologia , Tazobactam , Inibidores de beta-Lactamases , beta-Lactamases/metabolismo
16.
Microbios ; 67(271): 125-32, 1991.
Artigo em Inglês | MEDLINE | ID: mdl-1833612

RESUMO

It is not known how Mycobacterium leprae obtains energy for survival and growth in the host tissues; the organism does not grow in vitro. In the studies reported here, M. leprae incorporated labelled ATP, which was blocked by cyanide, unlabelled ATP or ADP, but not by adenosine or Pi. It seems that the organism takes up unhydrolysed ATP by an active transport process. The bacterium contained a membrane-bound, vanadate-sensitive E1 E2-ATPase (which creates a transmembrane potential driving transport of solutes into cells). The enzyme was not inhibited by N-ethylmaleimide, suggesting that it is not an F0F1-ATPase which catalyses ATP synthesis. Apparently, M. leprae derives energy-rich compounds from the host cell.


Assuntos
Adenosina Trifosfatases/metabolismo , Trifosfato de Adenosina/metabolismo , Mycobacterium leprae/enzimologia , Vanadatos/farmacologia , Difosfato de Adenosina/farmacologia , Adenosina Trifosfatases/antagonistas & inibidores , Trifosfato de Adenosina/farmacologia , Transporte Biológico Ativo/efeitos dos fármacos , Cianetos/farmacologia , Etilmaleimida/farmacologia , Concentração de Íons de Hidrogênio , Cinética , Mycobacterium leprae/efeitos dos fármacos , Mycobacterium leprae/metabolismo
18.
FEMS Microbiol Lett ; 58(1): 95-9, 1990 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-2118868

RESUMO

Water and soil samples were collected from natural habitats of the nine-banded armadillo and tested for the presence of acid-fast organisms by injection into the foot pads of experimental mice. Sixteen months post inoculation an acid-fast organism was isolated from the foot pad and spleen of one of the mice. The isolate exhibited diphenoloxidase activity as determined by its ability to convert D-3,4-dihydroxyphenylalanine to the corresponding quinone. The same organisms grown in vitro lacked detectable diphenoloxidase activity. However, diphenoloxidase activity was observed in acid-fast organisms harvested from spleen tissue of mice experimentally inoculated with a pure culture of the isolate. The environmental isolate was tentatively classed with the Mycobacterium avium-intracellulare complex.


Assuntos
Catecol Oxidase/metabolismo , Complexo Mycobacterium avium/isolamento & purificação , Microbiologia do Solo , Microbiologia da Água , Animais , Concentração de Íons de Hidrogênio , Camundongos , Camundongos Endogâmicos BALB C , Complexo Mycobacterium avium/enzimologia , Mycobacterium leprae/enzimologia
20.
J Basic Microbiol ; 29(1): 41-8, 1989.
Artigo em Inglês | MEDLINE | ID: mdl-2496221

RESUMO

This communication reports the association of changes in ultrastructure of Mycobacterium leprae with alterations in its permeability. To study morphologic changes of the organisms under different conditions (of temperature and exposure to NaOH and trypsin), ultrathin sections of the bacteria were cut and examined in an electron microscope. In the untreated bacilli and those washed with trypsin, the cytoplasmic membrane and the cell wall (peptidoglycan layer) remained intact; dapsone showed little effect on diphenoloxidase of the bacteria. M. leprae is unique among mycobacteria in possessing an unusual form of the enzyme diphenoloxidase. The antileprosy drug dapsone is a potent inhibitor of the enzyme, but it does not readily penetrate the bacteria where the cell envelope remains intact. The cell wall of M. leprae exposed to -80 degrees C or washed with NaOH was partially detached from the cell membrane; dapsone readily penetrated these organisms and inhibited the bacterial enzyme. In the above preparations, the cytoplasmic membrane appeared undamaged and the bacteria remained viable, as evidenced by multiplication in mouse foot pads. At 50 degrees C, the peptidoglycan layer became completely separated from the membrane and the cytoplasm was partially denatured. These organisms were permeable to dapsone, but were no longer viable. At 100 degrees C, the structural organization of the bacilli was completely destroyed, and of course, they lost their enzyme activity as well as viability. Evidently, the intact cell wall layer mediates the exclusion of dapsone from M. leprae, and there is no correlation between its viability and permeability. The ultrathin sections also reveal the internal organization and cytoplasmic inclusions of M. leprae, as never before seen.


Assuntos
Mycobacterium leprae/ultraestrutura , Catecol Oxidase/antagonistas & inibidores , Permeabilidade da Membrana Celular , Parede Celular/efeitos dos fármacos , Parede Celular/ultraestrutura , Dapsona/farmacologia , Microscopia Eletrônica , Mycobacterium leprae/efeitos dos fármacos , Mycobacterium leprae/enzimologia , Mycobacterium leprae/metabolismo , Hidróxido de Sódio/farmacologia , Temperatura , Tripsina/farmacologia
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