Different response to osteo-inductive agents in bone marrow- and periosteum-derived cell preparations.

Rabbit bone marrow- and periosteum-derived cells were cultured in medium containing dexamethasone, bone morphogenetic protein-2 (BMP2) or both. The response of bone marrow-derived cells, measured as alkaline phosphatase expression, depended on the stage of growth. In subconfluent cultures, BMP2 had a greater effect than dexamethasone and treatment with both further increased enzyme activity. In confluent cultures, the effect of dexamethasone was greater than that of BMP2 and, when used together, they had an additive effect. The mineral deposition observed in these cultures did not have the typical structure of bone nodules. For periosteum-derived cells, dexamethasone did not increase the expression of alkaline phosphatase, while BMP2 did; treatment with both was less effective than treatment with BMP2 alone. Typical bone nodules were observed in cultures of periosteum-derived cells treated with dexamethasone and BMP2. These findings indicate that either osteoprogenitor cells from these two sources are intrinsically different or else non-progenitor cells in the preparations directly or indirectly affect the responsiveness to osteo-inductive agents.

Monoclonal antibody against adult marrow-derived mesenchymal stem cells recognizes developing vasculature in embryonic human skin.

We have described previously a monoclonal antibody (SH2) that specifically recognizes undifferentiated mesenchymal progenitor cells isolated from adult human bone marrow. These cells, which we operationally refer to as mesenchymal stem cells, have the capacity to differentiate and form distinct mesenchymal tissues such as bone and cartilage when the isolated cells are placed in the appropriate in vivo or in vitro environment. We report here the partial biochemical characterization of the antigen recognized by the SH2 antibody. Metabolically radiolabelled adult marrow-derived mesenchymal stem cells in culture were extracted and immunoprecipitated with the SH2 antibody. The purified antigen migrated as a single band of 90 kDa after sodium dodecyl sulfate polyacrylamide gel electrophoresis was performed under reducing conditions. The SH2-immunoprecipitated protein exhibited a molecular weight band shift after removal of N-linked oligosaccharides. We investigated the expression of the SH2 antigen, along with the endothelial markers factor VIII-related antigen and Ulex europaeus I (UEA-I) lectin during specific developmental periods in human dermal embryogenesis and in the postnatal period through aged adults. Frozen sections of human embryonic, fetal, or postnatal skin ranging from 8 weeks estimated gestational age (EGA) through 84 years of age were immunostained or double immunolabelled with antibodies SH2, UEA-I, or factor VIII-related antigen followed by second antibodies with fluorescent markers. Positive cell surface reactivity with the SH2 antibody was seen in cells in the vascular plane in the earliest specimens (day 55 EGA) corresponding to the late cellular dermis period. During the period of the cellular to fibrous transition, in which the initiation of appendage development occurs, most SH2-reactive cells colocalized with vasculature markers UEA-I and factor VIII-related antigen, although there was a subset of cells recognized by SH2 antibody that did not colocalize with the endothelial markers. In contrast to the endothelial markers UEA-I and factor VIII-related antigen, in which the number of immunopositive cells became more prominent with age and maturation of the dermis, the frequency of cells that contained the SH2-reactive antigen diminished with age. The SH2 reactivity evident in embryonic, fetal, and early postnatal periods was not observed in human skin specimens taken from adults greater than 30 years old. These observations support the hypothesis that the SH2 antigen is a cell surface marker of developing microvasculature and may play a role in dermal embryogenesis and angiogenesis.

Osteochondrogenic potential of marrow mesenchymal progenitor cells exposed to TGF-beta 1 or PDGF-BB as assayed in vivo and in vitro.

Mesenchymal progenitors cells can be isolated from rat bone marrow and mitotically expanded in vitro. When these cells, which we operationally call mesenchymal stem cells (MSCs), are placed in an appropriate environment, they have the capacity to differentiate into bone and/or cartilage. This capacity is called osteochondrogenic potential. In this study, preconfluent MSCs were exposed in vitro to 5 ng/ml transforming growth factor-beta 1 (TGF-beta 1) or platelet-derived growth factor, isoform BB (PDGF-BB) for a pulse of 48 h and assayed for cell proliferation, alkaline phosphatase activity, and osteochondrogenic potential; untreated MSC's served as controls. In these cell culture conditions, TGF-beta 1 or PDGF-BB had similar effects on proliferation and alkaline phosphatase activity. Both growth factors increased cell proliferation and decreased alkaline phosphatase activity of MSCs. Sister cultures of TGF-beta 1- or PDGF-BB-treated MSCs and untreated MSCs were trypsinized. For each type of culture, the trypsinised MSCs were split in two parts: one part was replated in an osteogenic medium to assess its in vitro osteogenic potential, whereas the other part was seeded into porous calcium phosphate ceramics and implanted subcutaneously in syngeneic rats to assess its in vivo osteochondrogenic potential. PDGF-pretreated MSCs showed no difference in in vivo and in vitro osteochondrogenesis from that of control MSCs, while TGF-beta 1 pretreatment blocked the osteochondrogenic potential of MSCs when assayed in vitro for bone nodule formation. However, when tested in vivo, TGF-beta 1-pretreated MSCs were able to form bone and cartilage. These data show that measurements of proliferation and alkaline phosphatase activity of preconfluent MSCs immediately after exposure to growth factor were not predictive of their subsequent osteochondrogenic potential. Moreover, the variation of the osteochondrogenic potential of MSCs after exposure to growth factor was further modulated by the environment in which the MSCs were assayed.

Radioimmunodetection of rat and rabbit cartilage using a monoclonal antibody specific to link proteins.

Biodistribution analysis using [125I]Fab-6F3 specific to link proteins from human articular cartilage performed in rats by autoradiography showed a high concentration of radioactivity in all cartilaginous tissues. Preliminary immunoscintigraphic assays were performed in rabbits. Front and side view images of whole animals exhibited high uptake in cartilage tissue of the knee articulation, in the invertebral disk and the humeral head. This fixation was still detected 24 h post-injection, although high washout of radioactivity was observed.

Heterogeneity of neuropeptide Y immunoreactivity in patients with pheochromocytoma: influence on the diagnostic power of measuring plasma NPY using antisera with different specificities.

The nature of NPY-like immunoreactivity (NPY-LI) was investigated in plasma and tumour tissue of 17 pheochromocytoma patients by HPLC, gel filtration and isoelectric focusing using two radioimmunoassays (RIAs) directed against the C- and N-terminals of NPY respectively. The two RIAs gave similar results in pheochromocytomas: 86% of cases had higher NPY-LI concentrations than those found in normal adrenal glands and NPY-LI behaved like authentic human NPY during gel filtration and HPLC. Assessed by isoelectric focusing, NPY was found to be amidated in seven of nine tumours. Contrary to the findings obtained in tumours, the results of the two RIAs in plasma samples were not always concordant: compared to controls, elevated concentrations of NPY-LI were found in 86% of cases of pheochromocytomas using the C-terminally directed RIA and in 76% of cases using the N-terminally directed RIA. The results of HPLC and gel filtration of NPY-LI in plasma suggested that circulating C- and N-terminal NPY fragments account for the discrepancy between the results of the two RIAs. In conclusion, most pheochromocytomas contain large amounts of NPY-LI that behaved like authentic NPY by chromatographic analysis. On the contrary, circulating NPY-LI in some pheochromocytoma patients is heterogeneous with cleaved products which influence differently the power of the C- and N-terminally directed RIAs for the diagnosis of pheochromocytoma.

Biodistribution study of a monoclonal antibody specific to link proteins from human cartilage.

We report the use of monoclonal antibody 6F3 prepared against link proteins from human articular cartilage to elucidate the distribution of these glycoproteins within connective and other tissues. By immunohistochemical analysis, we showed that only the Fab fragment could reach the antigenic site in human articular sections. Cross-reactivity of the antibody 6F3 with link proteins purified from rat articular cartilage allowed us to carry out a biodistribution analysis in vivo in the rat. The time course of whole blood and plasma showed maximal activity 6 h after the 20 micrograms i.v. injection of [125I]Fab-6F3. Urinary excretion seems to be a high route of elimination. Moreover, we noticed no radioactive uptake across the blood-brain barrier. A significant fixation of labeled antibody Fab-6F3 was observed in noncartilaginous connective tissues such as aorta, skin and lung. As expected, specific and increased radioactivity was observed in all cartilage tissues, this increase was significantly higher 6 h after the [125I]Fab-6F3 injection than in the other connective tissues.

[Infection by the human immunodeficiency virus (HIV) in French Guyana. Dermato-venereologic problems]. / L'infection par le virus de l'immunodéficience humaine (VIH) en Guyane Française. Problèmes dermato-vénéréologiques.

Dermato-venereal manifestations in HIV infection and its severe evolution stage, AIDS, is of particular importance in tropical zones: We may be suspicious of the viral infection and consequently to request serologic tests to confirm it. We get an explanation of the virus transmission during heterosexual relations by the frequent occurrence and importance of the genital manifestations, leading to consider AIDS as a true sexually transmitted disease. Beside the classical opportunistic infections, the authors draw the attention to three types of manifestations: prurigo, already well known in Haïti and Africa capillary dystrophies, already reported in Haïti donovanosis that, because its epidemiological and etiopathological peculiarities, should be listed within the possible opportunistic infection if we take into consideration the regional pathological environment. In an other correction, syphilis, lepra and cutaneous leishmaniasis have to be carefully monitored, because they are capable to evaluate unexpectedly in some immunodepressive diathesis. Importance of dermato-venereal pathology in black people in tropical zone is explained by the weakness of cutaneous corneal stratum, immunologic disorders linked up to accumulated parasitic pathologies, socio-cultural life with a sexuality without complex.

PIP: Cutaneous manifestations of AIDS in the 1st 91 cases diagnosed in French Guiana between 1982-October 1987 included 40 cases of candidiasis, 29 of prurigo, 13 of herpes simplex, 5 of trichomoniasis, 7 of human papilloma virus, 3 of shingles, 3 of donovanoses, and 1 of Kaposi's sarcoma. There were also 7 cases of seborrheic dermatitis, 6 of capillary dystrophies, and 1 of leucoplasia. 26 of the 40 cases of candidiasis were buccal or buccopharyngeal and 14 were vaginal. Such infections are intense, chronic, and easy to diagnose. Local treatment with Nystatin or Amphotericin B in solution for buccal cases and with imidazole derivatives for vaginal cases should be supplemented with systemic medications such as ketoconazole. Most herpes simplex cases are type 2 genital infections which may be chronic and extensive. A perfusion of Aciclovir usually gives good results in 5 or 6 days. Shingles during AIDS often has nonthoracic localizations; involves itching, pain, and burning sensations; is recurrent, perhaps on the contralateral side; and may leave scars. Sensitivity to Aciclovir is less than for herpes simplex. Human papilloma virus lesions that are not too large are treated locally. Although tuberculosis is in 2nd place after candidiasis among opportunistic infections in AIDS patients in French Guiana. Only 2 cases of cutaneous tuberculosis were observed. 3 cases of Donovanosis due to Calymmatobacterium granulomatis were observed, with 2 cases with 1 couple. Chronic prurigo has been observed frequently in AIDS patients in Africa and Haiti. Along with asthenia, polyadenopathies, and shingles, it is often an early sign of AIDS. The pruritus becomes more and more intense and the only treatment providing some relief is local corticotherapy. The dermatovenereal signs of AIDS in tropical environments should raise suspicions of the disease in undiagnosed cases, and they also provide an explanation for the high rate of heterosexual transmission in individuals with various disorders involving genital lesions. Some dermatological disorders common in French Guiana have not been observed in AIDS patients to date.

[Human immunodeficiency virus (HIV) infection in mothers and infants in French Guyana. Epidemiologic study apropos of 44 women having conceived 55 children]. / L'infection par le virus de l'immunodéficience humaine (VIH) chez la mère et l'enfant en Guyane Française. Etude épidémiologique à propos de 44 femmes ayant eu 55 enfants.

The French Guyana is an Overseas French Department in South America, with 100,000 inhabitants among them are 20% of Haitian immigrants. At 31 December 1987, 103 AIDS cases have been recorded, 86% by heterosexual transmission. The first case dated May 1979 was retroactively diagnosed in an Haitian parturient, thank to her serum kept in the Pasteur institute of Cayenne. 44 women got their pregnancy during their HIV infection: 5 with clinical and biological evidence of AIDS, 7 developed AIDS after getting pregnant, and 10 out of these 12 women died. All of them were from a rather low social group and, generally, were not married. 43 are black (40 Haitians and 3 Guyanese Creoles), one is Indian and presented some psychic disorders. The mean age was 32 1/2 (from 15 to 51 years old). 55 babies were born: 12 developed AIDS (6 died during the first 15 months of their life); 14 are HIV +, 3 stillborns, 4 never tested and 22 developed negative reaction (with ELISA and Western-Blot) between 7.5 and 10 months of their life. Two Hutchinson's triads were observed. Prurigo is the most commonly skin manifestation observed.