Detection of human papillomavirus gene sequences in cell lines derived from laryngeal tumors.

The role of Human Papillomaviruses (HPV) in laryngeal carcinomas has been studied with conflicting results. To evaluate the etiologic relationship between HPV infection and epithelial malignancy of the larynx we studied five laryngeal carcinoma cell lines obtained from patients undergoing surgery for laryngeal tumors. The paraffin embedded biopsy samples of the original tumor and different passages of the new established cell lines were investigated by PCR with consensus primers specific for HPV DNA. The findings indicate that HPV infection is associated with some larynx carcinomas. The positive association has been enhanced when a method of enrichment of epithelial cells from fresh tumor samples was used. All tumor cells enriched smears were positive for HPV DNA not only by PCR but also by in situ hybridization (ISH). Investigated by PCR, different passages of larynx tumor cell lines maintained expression of HPV DNA. At subsequent passages ISH gives constantly no signals suggesting a minimal amount of viral harbored sequences. In one cell line propagated more than 60 population doublings, the chromosomal frequency distribution shifted from modal number 46 at the 5(th) passage to 63 at the 60(th) passage. The mechanisms by which persistent HPV infection maintains continuous cell proliferation were discussed.

Immunogenicity of inactivated vaccines prepared with parainfluenza viruses type 1 and type 3, administered experimentally by nasal or oral route.

The inactivated vaccines prepared with parainfluenza viruses type 1 and type 3, administered to mice by nasal or oral route, either as monovalent preparations in succession or as bivalent associated preparations according to the experimental models used, imparted a significant protection against the infection with the homologous active viruses. The routes of administration of vaccines, nasal and oral, which make equal demands upon the immune secretory serum and cell system, as well as the alternative of inactivated preparations for active virus vaccines are discussed.

Observations regarding the neurocytokaryological study of the experimental infection with herpes simplex virus type 1 and type 2 in the white mouse.

Mice distributed in groups were inoculated with the herpes virus simplex type 1 and type 2 strains in various dilutions (10(-1)-10(-7)), which had been maintained on primary human embryo cell cultures. The animals were killed and homogenates were prepared of their brains, which were used for testing the infective titre in vivo and for the isolation of nuclei, by means of the Hymer-Kuff technique. For the cytokaryological examinations, smears were prepared from the homogenate, which were stained by May-Grünwald-Giemsa and Mann methods. The cell and nucleus types were evaluated morphologically, descriptively, according to the size and structure. On the basis of three criteria the following groups were set up: large sized cells with bulky nuclei; middle sized cells with nuclei of a medium size; small sized cells with small nuclei and visible nucleoli; small sized cells with small, tachychromatic nuclei; cells exhibiting karyolysis. In both the infection with herpes virus simplex type 1 and type 2, the small sized cells with small tachychromatic nuclei represent the highest rate. Between the variants of the elaborated models no significant differences regarding the cell and nucleus types are recorded. On the whole, the cytokaryological picture does not exhibit notable differences, but rather resemblances with respect to the proportionality of the nucleus types in the experimental variants studied, in the case of herpes simplex virus type 1 and 2 induced infections.

Virological and pathomorphological aspects in experimental infections with some triassociated pneumotropic viruses in the white mouse.

Experimental infections were induced in white mice by intranasal administration of parainfluenza virus type 3, 739-2D strain, to which influenza virus A/Beijing, 353/89 (H3N2) strain, and respiratory syncytial virus, Long strain, were associated. The model was organized so as to obtain a triassociated infection, parainfluenza virus type 3 being inoculated the first and the other two viruses, in the following stages-II and III-, alternately. The infections were revealed by the presence of positive immunofluorescence reactions in the pulmonary tissue, of histological, histochemical and histoenzymatic lesions at the level of the respiratory apparatus, as well as of pathomorphological changes in other organs. At the pulmonary parenchyma level the inflammatory lesion had a 100% frequency. The severest pathomorphological picture was the diffuse, lymphohistiocytic and macrophagic bronchopneumonia. The cytoinfiltrate was characterized by a proportionality between lymphocytes and histiocytes and the lower but not insignificant presence of macrophages. The prevalent lesion was the thickening of interparietoalveolar septa, as a consequence of stasis hyperemia, oedema and lymphohistiomacrophagocytic infiltrate, to which dystrophic lesions, especially of biocytes, are often added. Different lesions are present in other organs, such as: in the liver-inflammatory, dystrophico-inflammatory and dystrophic lesions; in the kidney, pancreas, cerebellum and thymus--vascular changes, stasis hyperemia; in the spleen--hyperplasia of megakaryocytes. Generally, the type and severity of lesions vary from one viral-association-induced infection to another.

Testing of the in vivo therapeutic efficacy on the cutaneous herpes simplex infections of the autochthonous product 5% Acyclovir ointment.

The experimental results of the in vivo testing of an autochthonous pharmaceutical Acyclovir form prepared for the topical treatment of herpetic infections with a mucocutaneous location are shown in this paper. This testing on laboratory animals continues the in vivo performed investigations regarding the antiviral activity of this compound, which have proved that the efficacy of the inhibitory action exerted by the product on the Herpes simplex virus multiplication is comparable with the characteristics of the standard substance (Acyclovir-Zovirax Wellcome). By testing the therapeutic efficacy of the autochthonous Acyclovir preparation on an experimental model of cutaneous herpes infection in the newborn rat, it is demonstrated in a statistically significant manner that the product exerts a strong inhibitory action on the virus multiplication at the level of epidermis (proved by the lowering of virus production in the cutaneous tissue); the result is a drastic reduction of local herpes vesicles and of virus propagation in the neuraxis attended by the appearance of herpes meningo-encephalitis with a lethal course. The preparation is well tolerated (phenomena of local intolerance or remote toxicity were not observed). These in vivo positive results corroborated by those obtained in vitro complete the experimental argumentation necessary to support the proposal regarding the clinical trial of the product.

Some virological and pathomorphological aspects of the respiratory system in the experimental infection with respiratory syncytial virus associated with influenza virus, parainfluenza virus type 3 and adenovirus in the mouse.

Infections with respiratory syncytial virus Long strain, associated with influenza virus, A/Beijing 353/89 (H3N2) strain, parainfluenza virus type 3, 739-2D strain, and adenovirus type 3, were experimentally induced in white mice, causing histological, histochemical and histoenzymatic lesions at the respiratory system level, the severity of which exceeded the one observed in the controls infected with a single virus. The pathomorphological changes made up an inflammatory, predominantly infiltrative, lymphohistiocytic, then exudative and alterative picture. The severest and most frequent lesion was the diffuse interstitial, often peribronchiolovascular, bronchopneumonia, which might involve large parenchyma areas. Another highly frequent pulmonary lesion was the thickening of interalveolar septa, due to stasis hyperemia, oedema and the predominantly lymphocytic cytoinfiltrate. At the level of the extrapulmonary airways, the lesion present in all experimental models was the denudation of epithelium cilia. In the viral association in which influenza virus was included, an alteration, the hyalinosis of tunica media of the vessels, as well as of the Reisseisen's muscle, was also observed, in addition to the cytoinfiltrate; when the association was achieved with parainfluenza virus type 3, many macrophages and erythrocytes and a few fibroblasts appeared in the cytoinfiltrate, the alteration being the same as in the former model; when the association contained adenovirus, there appeared necrosis, abundant lymphocytes and lysis of the Reisseisen's muscle in the bronchopulmonary block. The associated infections were demonstrated by the presence of homologous serum antibodies and by positive IF reactions in the pulmonary tissue.

Experimental researches in infections with associated myxoviruses in the mouse.

Infections with influenza virus, A/Beijing 353/89 (H3N2) strain, to which there were associated parainfluenza virus type 3, 739-2D strain, adenovirus type 3, and respiratory syncytial virus Long strain, were experimentally induced in white mice. The experimental models were set up so as to permit the obtaining of an associated infection with three viruses, in which the influenza virus should be inoculated the first, the participation of the others being variable, according to their presence by alternation. The infections were detected by means of the presence of homologous serum antibodies, of positive immunofluorescence reactions in the pulmonary tissue, of the histological, histochemical and histoenzymatic lesions at the level of the respiratory system, as well as of pathomorphological changes in other organs. The severity of lesions varied from one to another infection produced by a viral association. At the level of the pulmonary parenchyma, the inflammatory lesion had a frequency of 100%. The severest pathomorphological picture characterized the diffuse interstitial lymphohistio-macrophagocytic bronchopneumonia. The bronchopulmonary block was marked by cytoinfiltrative processes, with a prevalence of lymphocytes in the infection with influenza virus + adenovirus + respiratory syncytial virus, but with a proportionality between lymphocytes and histiocytes in the other infections. The lesion of the highest incidence was the thickening of interalveolar septa, as a consequence of stasis hyperemia, oedema and lymphohistio-macrophagocytic cytoinfiltrate, sometimes associated with hyalinosis of tunica media of the blood vessels and of the Reisseisen's muscle. In other organs, particularly in the liver and kidney, vascular lesions, stasis hyperemia, inflammatory and dystrophico-inflammatory lesions were present; in the spleen, megakaryocyte hyperplasia was recorded at a significant rate in associated infections in which the adenovirus was present.

Experimental infections with some pneumotropic viruses in the mouse. Note II. Virological and pathomorphological aspects of experimental associated infections with parainfluenza virus, adenovirus and respiratory syncytial virus in the mouse.

Associated infections with long-strain parainfluenza virus type 3, adenovirus type 3 and respiratory syncytial virus were experimentally induced in white mice. They were pointed out by the appearance of homologous serum antibodies, positive IF reactions in the pulmonary tissue, and histological, histochemical and histoenzymatic lesions, which were more severe than those of controls infected with single virus of those mentioned above, since they involved wider areas and had a high frequency. The pathomorphological changes made up an inflammatory, exudative, alternative and predominantly infiltrative lymphohistiocytic picture, mainly pulmonary (diffuse interstitial bronchopneumonia, sometimes with a peribronchovascular location, of a cuff-like aspect), hepatic, renal and cardiac, but dystrophic processes were also present--hyalinosis of tunica media in the lung, hepatocyte cytoplasm vacuolizations on areas of various sizes of lobuli--, especially in associated infections with parainfluenza virus type 3 and adenovirus type 3. Megakaryocytic hyperplasia in the spleen, present in all the experimental models, was also described. Generally, the modified structural aspects made up and pointed out a complex pathological process.

Testing of some pharmacological characteristics of the autochthonous product Acyclovir.

In the present work, the preliminary results of a study regarding the pharmacokinetic properties of the autochthonous synthetic drug Acyclovir are shown. The in vitro and in vivo toxicologic test demonstrates a low toxicity of this product, the drug being well tolerated both by the laboratory animal and by the fibroblasts of the human embryo. The specific antiviral action of the product on the multiplication in vitro of the Herpes simplex virus types 1 and 2, appreciated by the lowering capacity of the infection titre and by the determination of the inhibitory concentration 50% (IC50), is significant and comparable with previous results obtained by the authors by testing the drug Zovirax of the Welcome firm.

Experimental infections with some pneumotropic viruses in the mouse. Note I. Virological and pathomorphological aspects of the associated infections with influenza virus, adenovirus and respiratory syncytial virus.

In white mice, the associated infections with the influenza virus type A(H3N2), the adenovirus type 3 and the long strain of the respiratory syncytial virus, detected by the appearance of homologous serum antibodies and by positive I.F. reactions in pulmonary tissue, induce histological, histochemical and histo-enzymatic lesions, the severity of which exceeds that of the lesions in animals infected with only one of the mentioned viruses. In comparison with the morphological picture in animals inoculated with a single virus, expressed by inflammatory, infiltrative, lymphohistiocytic pulmonary (infiltrative, interstitial, diffuse, sometimes "cuffing", peribronchial and perivascular broncho-pneumonias), hepatic and renal processes, in the case of associate infections pathomorphological changes appear on wider surfaces and with a high frequency, manifested also by hyalinosis of the vascular media in the lung and the spleen, vacuolization of the cytoplasm of hepatocytes, hypertrophy of the lymphoid follicles in the spleen and the lymph nodes (in associated infection due to the influenza virus+adenovirus and vice versa) or, on the contrary, hypotrophy of the lymphoid follicles in the spleen and lymph nodes (in the associated infection due to the influenza virus+respiratory syncytial virus and vice versa).

[Pathomorphologic features in several phlebopathies associated with virus, rickettsia, chlamydia, and mycoplasma infections]. / Aspects pathomorphologiques dans quelques phlébopathies associées à des infections chroniques à virus, à rickettsies, à chlamydies et à mycoplasmes.

Seventy-six venous specimens were examined by indirect immunofluorescence and histological techniques. 80.26% out of them gave negative I.F. reactions. The following causal agents were found on the I.F. positive preparations: Rickettsia burneti, Chlamydia, Mycoplasma, parainfluenza type 3 and adenoviruses, alone or associated. The most frequent lesion noted in I.F. positive specimens was of the inflammatory-dystrophic type. In I.F. negative preparations the dominant lesion was the hyalinosis of the tunica media.

[Morphologic changes following experimental influenza virus infection and the therapeutic administration of flavonoids in aqueous extract]. / Modifications morphologiques consécutives à l'infection expérimentale à virus de la grippe et à l'administration thérapeutique de flavonoïdes en extrait aqueux.

Influenza virus strain A/PR8/34(H1N1) and flavonoid aqueous extract with therapeutical function were administered by intranasal route. Pathological examinations were done three and ten days after administration to evaluate the morphological modifications established in the lung parenchyma. Results showed that therapeutical administration of flavonoids had good effects on interstitial bronchopneumonia evolution till complete recovery. Flavonoid aqueous extract administered by intranasal route was well tolerated by the organism.

[The pathomorphological aspects of arteriopathies associated with viral and microbial infections]. / Quelques aspects de pathomorphologie des artériopathies associées à des infections à virus et inframicrobiennes.

Presence of Mycoplasma, Chlamydia, Rickettsia burnetii, influenza type B, herpes and adenoviruses was detected by immunofluorescence in artery fragments collected from patients with arteriopathy. Pathohistological examination of the samples revealed inflammatory--lympho-histiocytic infiltration--and mostly dystrophic alterations. Arteriopathies were localised mostly at the pelvic limb level, especially on femoral artery. Their incidence was statistically significant.

[Features of the patho-morphology of arteriopathies associated with viral and infra-microbial infections]. / Quelques aspects de pathomorphologie des artériopathies associées à des infections à virus et inframicrobiennes.

Presence of Mycoplasma Chlamydia, Rickettsia burnetii, influenza type B, herpes and adenoviruses was detected by immunofluorescence in artery fragments collected from patients with arteriopathy. Pathohistological examination of the samples revealed inflammatory--lympho-histiocytic infiltration--and mostly dystrophic alterations. Arteriopathies were localised mostly at the pelvic limb level, especially on femoral artery. Their incidence was statistically significant.

[New pathogenetic aspects of the relations between viral and microbial infections and peripheral vasculopathies]. / Quelques nouveaux aspects pathogéniques des relations entre certaines infections à virus et inframicrobiennes et des vasculopathies périphériques.

The immunofluorescence technique using antisera against some viruses and inframicrobes allowed the detection of pathogens in altered vascular tissues (arteritis and phlebitis-phlebectasia). Pathomorphological aspects and some dehydrogenase activities in these patients were also investigated.

[A morphological study of pneumopathies caused by associated para-influenza and influenza viruses]. / Etude morphologique des pneumopathies provoquées par les virus paragrippal et grippal associés.

Histological examination of lungs collected from mice inoculated with influenza A/PR 8/34 (H0N0) and/or parainfluenza type 3 viruses revealed the existence of lymphohistiocytic interstitial bronchopneumonia lesions. Necrosis zones were found in the lungs of influenza virus infected animals. Associated infection by the two viruses induced also the fibroblast and fibrocyte proliferation, at same average ratios.