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Interaction of Clostridium difficile toxin a with cultured cells
Ceil Biology
IIITERACTIOY OF cu)sloIDIuII QIPPICILE TDXIY A WITH UJLTUReD CEUS
Fv25
Carla Fiorentini, Silvia Paradini, Narina Giuliano, 1stituto Paola Rastrontonio, Gianfranco Donelli. Superiore di Sanita’, Roma, Italy. The anaerobic bacterium Clomtridiuw difficile produces at least tro protein toxins, toxins A and implicated in the pathogeneais of antibioticB. associated paeudom?nbrenous colitis. Toxin A is an enterotoxin vhich in addition is slightly cytotoxic vhile toxin B is a potent cytotoxin lacking enterotoxic activity. Toxin A induces a eerries of cytopathological changes in cell cultures, mainly conaiin cell retraction and rounding and in the sting displacement of the nucleus tovards one pole of the cell .These changes appeared to be dependent on the cytoakeletal organization. In fact, miorofilaeent system seemed to be modified in the early st@pa of the intoxication process before the occurring of cell retraction. Hicrotubule netvork integrity and function appeared to be necessary for the nuclear displaceaent in that the ricrotubule depolymeriting agent demecolcin completely blocked the polarization of the nucleus. The intermediate filament vimentin vaa condensed near the polarized nucleus in rounded cells. lloreover, the cytopathic effect vas completely blocked by i1 the carboxylic ionophore monensin, when added to the cells together with the toxin and by ii) the serine protease inhibitor chymostatin vhich appeared to be protective upon addition also long after the end of the binding step. an acidic compartment and the Thus, action of a eerine proteaae seened to be essential for the activity of toxin A.
EFFECTS
OF PETROLEUM HYDROCARBONS ON THE OXIOATIVE METABOLISM OF DIGESTIVE GLAND CELLS OF MUSSELS. Mlren P. Cajaravllle, Joso A. Uranga, lonan Marlgomar and Eduardo Angulo. Zaol.-Hlatol. Lab., Biol. ~41. et4 zlmtzls
P727
Yorlol.
Saila,
Zlrntrl
Fak.,
E.H.U.,
644
P.K.,
48080-Bllbo,
Spaln.
Recent studies have related the toxicity of petroleum hydrocarbons to the metabolic production of toxic free oxyradicals through the mixed function oxygenase system. Then, enzymes comprising the endogenous antioxidant defense system, such as catalase, can be expected to be stimulated. We report here the effects of the Water Accomodated Fraction (WAF) of different oils on the oxidative metabolfsm of mussels Mytilus galloprovincialis Lmk. Individuals were exposed in replicated series for three months to different dilutions of the WAF of two crude oils and a commercial lubricant oil. Catalase activity was demonstrated in cryostat sections of the digestive gland by a novel histochemical procedure (1) in mussels sampled after 21, 49 and 91 days exposure. Reaction product was quantified in a Zeiss cytospectrophotometer at 490 nm. The mean absorbance was significantly increased in mussels exposed to the three petroleum hydrocarbons tested although a dose-dependent response was not always evidenced. The analysis of variance indicates exposure-time and the exposurethat both the concentration have a significant effect on the catalase activity of mussel digestive gland cells. Changes in glutamyl transpeptidase localization and activity were also quantified after histocbemical demonstration by the procedure of Rutenburg CI al . (2). m: This study has been partially financied by the Basque Government through a research fellowship to M.P.C. (1) Cajaraville M.P., Uranga J.A. & Angulo E. HISTOCHRMISTRY, submitted. (2) Rutenburg A.M., Kim H., Fischbein J.W., Hawker Wasserkrug ILL. & Seligman A.M. 1969. J. J.S., HISTOCERM. CY’TOCAEM., 17, 517-526.
International
P726
Reports,
Vol. 14, Abstracts
Supplement
1990
OF CHROMIlJM(V1) ON MAIZE ROOTS CYCLE AND NUCLEAR STRUCTURE
THE EFFECT II. CELL
Rita Musetti+,Maria Biology, University of Milan; University of Parma, ITALY. Seedlings of two maize inbred lines, 33.16 and 868, characterized respectively by high and low chromium tolerance, were treated in hydroponic solution with Sppm Cr(V1). At different day intervals, root apices were fixed in formaldehyde; nuclei were extracted from the meristematic tissue and stained with DAPI for fluorescence microscopy and flow cytometric analysis. Root samples were also processed for electron microscopy (TEM). In the 868 line, Cr has a marked effect on cell cycle and nuclear morphology. The percentage of cells in S was reduced from 35 to S-K%, with an arrest of cells mainly in Gl. At fluorescence microscopy, many nuclei presented a very large nucleolus and a diffuse fluorescence, besides slterations in size and shape. These features seemed to be due to alterations of the nuclear envelope, as suggested by TEM observations, On the contrary, in the tolerant line 33.16, after 7 days of treatment the nuclei were well conserved and a high proportion of S cells persisted throughMarisa Levi’, Maria ODept.of A. Favali+. +Instit.ut of Botany,
G.Corradi+,
out the treatment. These data are in accordance tion of root growth induced 868 line, in which also
led a higher level.
with the bycrtreatment
strong
inhibi-
in had
atomic dosorption accumulation of the metal at root
the revea-
p728
LEAD TOXIiJITY ON THE COCYTESOF THE T!&EOST TILAPIA nilotica bnnabelle h. Herrera,June Anne Luzares. Institute l f Eiolegy,C*llege of Science, ihiiversity of the Yhilippines,Dilman,?uesen City, Philippines. This is the first study on the effects of chronic exp4surc l f T. niletica_fry to sublethal lead.After acclimation for l ne week,juvenile fish were distributed equally to three aquaria& fourth aquarium? containing fish of the same number was used as the ccntrbl. Water was changed every two days and lead level was adjusted accrrdingly.lfter l ne menth,ovarFes were dissected l t.tt and processed for light and electren micrescepe analysis.Ultraatructural damage was rbserved such as: discontinuity in the mrmbranes,dilatirn of the ER,
mitrchondria braner of
and cell
lysosrmes,rupt:1:e of meman& ciumgini: of
l ryanelles
chrematin. These are cridencas that lead ts the recytes ef Tilapia nilstica, dy species l f ccenemic impbrtance i hili?pines.
ie
texic
a harin
the
IIITERACTIOY OF cu)sloIDIuII QIPPICILE TDXIY A WITH UJLTUReD CEUS
Fv25
Carla Fiorentini, Silvia Paradini, Narina Giuliano, 1stituto Paola Rastrontonio, Gianfranco Donelli. Superiore di Sanita’, Roma, Italy. The anaerobic bacterium Clomtridiuw difficile produces at least tro protein toxins, toxins A and implicated in the pathogeneais of antibioticB. associated paeudom?nbrenous colitis. Toxin A is an enterotoxin vhich in addition is slightly cytotoxic vhile toxin B is a potent cytotoxin lacking enterotoxic activity. Toxin A induces a eerries of cytopathological changes in cell cultures, mainly conaiin cell retraction and rounding and in the sting displacement of the nucleus tovards one pole of the cell .These changes appeared to be dependent on the cytoakeletal organization. In fact, miorofilaeent system seemed to be modified in the early st@pa of the intoxication process before the occurring of cell retraction. Hicrotubule netvork integrity and function appeared to be necessary for the nuclear displaceaent in that the ricrotubule depolymeriting agent demecolcin completely blocked the polarization of the nucleus. The intermediate filament vimentin vaa condensed near the polarized nucleus in rounded cells. lloreover, the cytopathic effect vas completely blocked by i1 the carboxylic ionophore monensin, when added to the cells together with the toxin and by ii) the serine protease inhibitor chymostatin vhich appeared to be protective upon addition also long after the end of the binding step. an acidic compartment and the Thus, action of a eerine proteaae seened to be essential for the activity of toxin A.
EFFECTS
OF PETROLEUM HYDROCARBONS ON THE OXIOATIVE METABOLISM OF DIGESTIVE GLAND CELLS OF MUSSELS. Mlren P. Cajaravllle, Joso A. Uranga, lonan Marlgomar and Eduardo Angulo. Zaol.-Hlatol. Lab., Biol. ~41. et4 zlmtzls
P727
Yorlol.
Saila,
Zlrntrl
Fak.,
E.H.U.,
644
P.K.,
48080-Bllbo,
Spaln.
Recent studies have related the toxicity of petroleum hydrocarbons to the metabolic production of toxic free oxyradicals through the mixed function oxygenase system. Then, enzymes comprising the endogenous antioxidant defense system, such as catalase, can be expected to be stimulated. We report here the effects of the Water Accomodated Fraction (WAF) of different oils on the oxidative metabolfsm of mussels Mytilus galloprovincialis Lmk. Individuals were exposed in replicated series for three months to different dilutions of the WAF of two crude oils and a commercial lubricant oil. Catalase activity was demonstrated in cryostat sections of the digestive gland by a novel histochemical procedure (1) in mussels sampled after 21, 49 and 91 days exposure. Reaction product was quantified in a Zeiss cytospectrophotometer at 490 nm. The mean absorbance was significantly increased in mussels exposed to the three petroleum hydrocarbons tested although a dose-dependent response was not always evidenced. The analysis of variance indicates exposure-time and the exposurethat both the concentration have a significant effect on the catalase activity of mussel digestive gland cells. Changes in glutamyl transpeptidase localization and activity were also quantified after histocbemical demonstration by the procedure of Rutenburg CI al . (2). m: This study has been partially financied by the Basque Government through a research fellowship to M.P.C. (1) Cajaraville M.P., Uranga J.A. & Angulo E. HISTOCHRMISTRY, submitted. (2) Rutenburg A.M., Kim H., Fischbein J.W., Hawker Wasserkrug ILL. & Seligman A.M. 1969. J. J.S., HISTOCERM. CY’TOCAEM., 17, 517-526.
International
P726
Reports,
Vol. 14, Abstracts
Supplement
1990
OF CHROMIlJM(V1) ON MAIZE ROOTS CYCLE AND NUCLEAR STRUCTURE
THE EFFECT II. CELL
Rita Musetti+,Maria Biology, University of Milan; University of Parma, ITALY. Seedlings of two maize inbred lines, 33.16 and 868, characterized respectively by high and low chromium tolerance, were treated in hydroponic solution with Sppm Cr(V1). At different day intervals, root apices were fixed in formaldehyde; nuclei were extracted from the meristematic tissue and stained with DAPI for fluorescence microscopy and flow cytometric analysis. Root samples were also processed for electron microscopy (TEM). In the 868 line, Cr has a marked effect on cell cycle and nuclear morphology. The percentage of cells in S was reduced from 35 to S-K%, with an arrest of cells mainly in Gl. At fluorescence microscopy, many nuclei presented a very large nucleolus and a diffuse fluorescence, besides slterations in size and shape. These features seemed to be due to alterations of the nuclear envelope, as suggested by TEM observations, On the contrary, in the tolerant line 33.16, after 7 days of treatment the nuclei were well conserved and a high proportion of S cells persisted throughMarisa Levi’, Maria ODept.of A. Favali+. +Instit.ut of Botany,
G.Corradi+,
out the treatment. These data are in accordance tion of root growth induced 868 line, in which also
led a higher level.
with the bycrtreatment
strong
inhibi-
in had
atomic dosorption accumulation of the metal at root
the revea-
p728
LEAD TOXIiJITY ON THE COCYTESOF THE T!&EOST TILAPIA nilotica bnnabelle h. Herrera,June Anne Luzares. Institute l f Eiolegy,C*llege of Science, ihiiversity of the Yhilippines,Dilman,?uesen City, Philippines. This is the first study on the effects of chronic exp4surc l f T. niletica_fry to sublethal lead.After acclimation for l ne week,juvenile fish were distributed equally to three aquaria& fourth aquarium? containing fish of the same number was used as the ccntrbl. Water was changed every two days and lead level was adjusted accrrdingly.lfter l ne menth,ovarFes were dissected l t.tt and processed for light and electren micrescepe analysis.Ultraatructural damage was rbserved such as: discontinuity in the mrmbranes,dilatirn of the ER,
mitrchondria braner of
and cell
lysosrmes,rupt:1:e of meman& ciumgini: of
l ryanelles
chrematin. These are cridencas that lead ts the recytes ef Tilapia nilstica, dy species l f ccenemic impbrtance i hili?pines.
ie
texic
a harin
the