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Biomedical subjects

L Germain

Publications and source records attributed to L Germain.

At least 73 records · Page 4Linked to original sources
PubMed1993

Neuronal activity in the motor and premotor cortices before and after learning the associations between auditory stimuli and motor responses.

In order to assess the effect of learning a conditional motor task on set-related cells of the precentral cortex we recorded a total of 228 task-related cells in awake, behaving primate. A first sample of 54 set-related cells was recorded while the monkey was performing at 54.7 +/- 7.8% and a second sample of 119 set-related cells was recorded at the same stereotaxic coordinates while he was performing at 77.2 +/- 8.7%. After the monkey had learned the association between an auditory signal and a motor response we found a significant increase in the proportion and activity of directional set-related cells in the premotor cortex. Furthermore the proportion of short-latency set-related cells was found to increase in this area. None of those changes were observed in the motor cortex.

Acoustic Stimulation↗
PubMed1993

Role and innocuity of Tisseel, a tissue glue, in the grafting process and in vivo evolution of human cultured epidermis.

Cultured epidermal sheets are currently used for burn wound treatment but reported results on graft take are variable. This study was designed to evaluate the role and influence of Tisseel, a fibrin glue, in the take of cultured human epidermal sheets in an athymic mouse model. On days 4, 10 and 21 post-grafting, histology, electron microscopy and immunofluorescence staining confirmed the presence of a human epithelium and the development of a basement membrane. Tisseel was detectable on day 4 only, but overall treated and untreated grafts were similar. The use of Tisseel enhanced the mechanical stability of these fragile grafts, increased the percentage of graft take, and its innocuity on the in vivo evolution of cultured epidermal sheets was demonstrated. For these reasons, we think that Tisseel may be advantageous in a clinical setting.

Animals↗
PubMed1993

Improvement of human keratinocyte isolation and culture using thermolysin.

We propose a modification of the conventional keratinocyte isolation method which has shown a significant improvement in the purity, colony forming efficiency (c.f.e.) and growth capacity of the isolated epidermal cell population. This method utilized thermolysin since it selectively digests the dermo-epidermal junction. Following separation from the dermis, the epidermis was digested with trypsin to obtain a single cell suspension. Compared with the conventional procedure, this isolation method was shorter and resulted in (i) cells displaying a higher colony forming efficiency, (ii) cells reaching confluence 1-3 days earlier, (iii) cells not contaminated by fibroblasts, (iv) a cell population containing all the basal layer keratinocytes. These cells were suitable for the establishment of primary cultures and could be subcultured. Such cell populations should be advantageous in studies of epithelial-mesenchymal interactions in which keratinocyte populations, free of fibroblasts, are desirable. In the treatment of extensively burned patients using cultured epidermal sheets, the main problem remains the time required for their production. Thus, the absence of fibroblast overgrowth of the keratinocyte cultures and the significantly reduced time to obtain confluent cultures and epidermal sheets with our method have very important implications for the treatment of large burn wounds.

Adolescent↗
PubMed1993

In vitro construction of a human blood vessel from cultured vascular cells: a morphologic study.

PURPOSE: The purpose of this study was to create a tubular vascular model exclusively made of human cells and collagen. METHODS: The blood vessel equivalent was constructed with the three following human cell types: vascular smooth muscle cells, endothelial cells, and fibroblasts. A tissuelike structure was obtained from the contraction of a tubular collagen gel (human origin) by vascular smooth muscle cells, which created a media-like structure. An adventitia-like tissue was added around the media-like structure by embedding fibroblasts into a collagen gel. An endothelium was established within the tubular structure after intraluminal cell seeding. RESULTS: Cell orientation and gel contraction were followed up over time. Vascular smooth muscle cells developed a complex tridimensional network and were oriented in a circular fashion around the tube's axis. In contrast, fibroblasts were randomly oriented. A viable, homogeneous, and well-characterized endothelium was observed. These endothelial cells showed a slightly elongated structure and were oriented parallel to this vascular equivalent axis. CONCLUSION: An in vitro tridimensional vascular model that exhibits some phenotypic characteristics of in vivo vascular cells could be useful in the study of events that lead to atherosclerotic plaque formations.

Cells, Cultured↗
PubMed1993

Cultured epithelium allografts: Langerhans cell and Thy-1+ dendritic epidermal cell depletion effects on allograft rejection.

The effects of in vitro dendritic cell (DC) depletion on the survival of epidermal sheet allografts were studied in a murine model. Newborn (1-3 days old) mouse skin was used. Langerhans cell (LC) and Thy-1+ dendritic epidermal cell (Thy-1+ DEC) depletion was achieved using: (1) a prolonged culture period (7 days), or (2) the anti-IA and anti-Thy-1.2 mAbs followed by complement treatment. DC (LC and Thy-1+ DEC) depletion was assessed on sheets and cultured cell suspensions by an indirect immunofluorescence procedure. They showed that, after 7 days of culture or after the antibody-complement treatment, epidermal cultures were depleted of LC and Thy-1+ DEC. Cultured sheets were grafted onto the muscle of H-2-incompatible recipients. The control experiments were: (1) full epidermis and DC undepleted sheet allografts, and (2) DC depleted and undepleted sheet isografts. The full epidermis was totally rejected after 9 days. However, no rejection sign was ever seen in any of the isografts. The in vitro produced and allografted epithelia did not show any necrotic sign until the 11th day postgrafting. However, on days 12-13, the DC depleted allograft's color changed from pink to brown. On days 14-16, degradation of the allografts resulted in a complete denudation of the underlying muscle. Immunohistological analysis of the allografts revealed the presence of a monocyte and lymphocyte infiltration starting from the 11th day postgrafting, with the presence of polymorphonuclear leukocytes on day 14. These results suggest that LC and Thy-1+ DEC depletions were not sufficient to prevent allograft rejection.

Animals↗
PubMed1992

Optimization of murine keratinocyte culture for the production of graftable epidermal sheets.

The aim of the present study was to optimize murine epidermal cell cultures in order to obtain graftable sheets. Newborn (1-3 days old) Balb/c mice skin were used to optimize culture media and plating cell concentration, then epidermal sheet production, and grafting. Epidermal cells were plated at various concentrations in different culture media containing low (0.1 mM) or high (greater than 1 mM) Ca2+ levels. After a 3 day culture at the 10(4) cells/cm2 plating cell concentration, the percentage of differentiated cells was more than 80% in the high Ca2+ culture medium and less than 50% with bulky cells in the low Ca2+ culture medium. Under these conditions confluence was not obtained. At the 10(5) cells/cm2 seeding inoculum, the percentage of confluence increased to 95-100% during the first 72 h of culture in both high and low Ca2+ culture media. Three-day-old culture showed stratified multilayer epidermal sheets in the high calcium medium, and monolayer epidermal sheets were present in the low calcium medium after seeding keratinocytes in fibronectin precoated flasks. Seven days after plating, post confluent cultures were composed of a high percentage of differentiated cells (90%) with an increase in shedding cells in the medium. Considering the above morphological observations, sheets obtained with 10(5) cells/cm2 in MCDB-153 (A), DME-HAM (B) or GMEM (C) media after 3 days in culture were grafted. Twenty days after grafting, histological analysis of biopsies showed an epidermal structure and organization comparable to normal murine epidermis without hair follicles. Epidermal transplants showed a complete basement membrane, hemidesmosomes, and tonofilament bundles. Sheets obtained after seven day culture in all media showed lower coverage of the wound bed. These studies point out the importance of the plating cell and Ca2+ concentrations, and the culture time for murine keratinocyte confluence and differentiation to obtain graftable epidermal sheets.

Animals↗
PubMed1992

Peripheral anchorage of dermal equivalents.

Human fibroblasts can induce collagen gel contraction with different kinetics depending on the number of cells and on the collagen concentration within this lattice, which has been considered as a dermal equivalent. Skin equivalent is a combined culture of dermo-epidermal layers which may be of therapeutic value in the treatment of burn patients. However, the current production of the dermal equivalent component gives results that present many drawbacks for their eventual clinical use as a first step in obtaining a skin equivalent. These include: (i) final surfaces which are very small; less than 20% of the initial size (ii) excessive thickness which may hamper successful graft take (iii) fibroblasts that do not have an arrangement comparable with normal dermal tissue. We propose, as a solution to these problems, the utilization of a 5-mm-wide fibre-glass filter ring peripherally attached to the surface of the Petri dishes to prevent inordinate contraction while the fibroblasts reorganize the collagen gel. Using this technique the initial surface was preserved and the dermal equivalent contracted only in thickness. Histological analysis of these anchored equivalents confirmed an alignment of fibroblasts and collagen fibres resembling normal dermal tissue. We consider this method useful in the development of dermo-epidermal sheets for clinical purposes.

Bioprosthesis↗
PubMed1992

A descriptive study of epilepsy in the district of El Salvador, Chile, 1984-1988.

To describe the epidemiological characteristics of epilepsy in a northern area of Chile, an investigation was conducted in four localities in the province of Copiapó (population of 17,694). Based on 314 cases of active epilepsy, the prevalence per 1000 at June 30, 1988 was 17.7. The average annual incidence for the period 1984-1988 was 113 per 100,000. Partial seizures were the most frequent type of seizure diagnosed (54.1%). Antecedents considered as possible etiological factors were found in 29.9% of cases. According to age of onset, 64.6% had their first attack before 15 years. Middle and low socioeconomic classes had higher prevalence rates of epilepsy. We compare our results with previous Latin-American studies.

Adolescent↗
PubMed1992

Influence of dermal equivalent maturation on the development of a cultured skin equivalent.

Histologic and immunofluorescence methods were used to analyse the presence of fibronectin, chondroitin-4-sulphate and chondroitin-6-sulphate, type III and IV collagens, laminin, and keratins to assess the maturation level of cultured dermal and skin equivalents. In a first phase, fibroblasts in monolayer culture were compared with dermal equivalents in which fibroblasts are embedded in a type I collagen gel. Different fluorescent patterns were observed depending on the culture system used. A sequential appearance of macromolecules was noticed in dermal equivalents. Fibronectin was first detected after 4 days of culture, whereas chondroitin-4-sulphate and chondroitin-6-sulphate and type III collagen were present after 7 days. In contrast, all three macromolecules were detected at 24 h of culture in fibroblastic monolayer cultures. In a second phase, the quality of our skin equivalents was evaluated according to the seeding time of epidermal cells upon dermal equivalents (1, 4, or 7 days). A satisfactory stratification was obtained when keratinocytes were seeded after 4 and 7 days of dermal equivalent culture. Laminin and fibronectin were detected at the dermo-epidermal junction, but type IV collagen was absent. Various keratins, as detected by the AE1, AE2, and AE3 antibodies, were present in the epidermal layer. Following keratinocyte confluence, a change in the organization pattern of type III collagen in the dermal fraction of the skin equivalent was also noticed. Our comparative results show that seeding of epidermal cells on a more mature dermal equivalent leads to improved differentiation status of the epidermal layer.

Cell Division↗
PubMed1992

Comparison of immunocytochemical and Holzer's methods for detection of acute and chronic gliosis in human postmortem material.

Immunocytochemical staining for glial fibrillary acidic protein (GFAP) allows more specific identification of astrocytes and their processes than classical histochemical techniques and has therefore recently been used by some investigators to quantify gliosis. However, although the immunocytochemical method is superior for delineation of reactive astrocytes, the examples presented here and previous work by others demonstrate that chronic fibrillary gliosis may be best detected by Holzer's method and not by GFAP immunocytochemistry. The authors' studies indicate that if, as in a recent study of gliosis in schizophrenia, computer-assisted densitometry is to be used to measure gliosis, the immunoperoxidase method may not be a sensitive technique to demonstrate glial changes in human postmortem material.

Brain↗
PubMed1992

[Murine surgical model for the study of cultured grafts].

The objective of the study was to establish an animal model for in vivo studies of cultured cutaneous equivalents. The model on athymic mice that we already described (López-Valle C.A. et al., Plast Reconstr Surg, 1992, 89, 139-143) satisfied the criteria of immobilization of the recipient site and physical stability of the graft, but still allowing complete movement freedom of the animal used. Nevertheless, this technique encountered two long term weaknesses; a) a significant grafted surface reduction caused by the wound contraction and b) the absence of a physical barrier between human and murin keratinocytes. We propose some modifications to this technique to correct both problems. Moreover, the implantation of polypropylene, instead of glass pellets, to generate granulation tissue on the recipient bed when needed, constitutes an easier method for both the surgeon and the animal. Finally, in order to minimize wound care, some modifications were made to the rodent cages. Immunohistological analyses of the biopsies, 21 days post-grafting, revealed a continuous basal membrane. This animal model allows in vivo studies on the behavior, cicatrization and immunology of human cultured skin equivalents.

Animals↗
PubMed1991

Cellular basis for the elevated gallium-67 computed lung index in a rheumatoid lung patient.

A patient with high levels of serum rheumatoid factor and an open lung biopsy which showed high-grade interstitial pneumonia with large numbers of lymphocytes and plasmocytes had intense gallium uptake in the lungs. Lymphocytes and/or plasmocytes might be responsible for the gallium uptake even though neutrophils are usually credited with high-level uptake. Differential cell counts demonstrated plasmocyte and lymphocyte preponderance, but neutrophil paucity. In vitro cell cultures of purified neutrophils, monocytes, leukemic plasmocytes, and resting and stimulated lymphocytes with 67Ga showed that plasmocytes take up comparatively low levels of 67Ga, but that activated lymphocytes take up levels that approach neutrophils. It is probable that both rheumatoid lung plasmocytes and activated lymphocytes are responsible for the pulmonary 67Ga concentration in this patient.

Aged↗
PubMed1990

Influence of initial collagen and cellular concentrations on the final surface area of dermal and skin equivalents: a Box-Behnken analysis.

Our laboratory has been involved in finding optimal conditions for producing dermal and skin equivalents. As an original approach, a Box-Behnken experimental design was used to study the effects of the initial collagen and fibroblast concentrations and the initial gel thickness on the contraction of dermal and skin equivalents. The final surface area of dermal equivalent varied significantly with the initial concentration of collagen and fibroblast, whereas the initial thickness of gel had no appreciable effect on the contraction of the dermal equivalent. When keratinocytes were grown on these dermal equivalents they produced a very severe contraction, to an extent that all skin equivalents had a similar final surface area. This severe contraction was independent of collagen and fibroblast concentrations. Models for the prediction of the final percentage contraction of dermal and skin equivalents as a function of the initial concentration of collagen, the logarithm of fibroblast concentration, and the initial gel thickness were obtained and analyzed. Keratinocytes grown at the lowest seeding density did not contract the equivalents. However, histologic analysis has shown an incomplete coverage by these cells of the equivalents. The extensive contraction of the skin equivalent presenting adequate morphology is a major drawback toward its clinical utilization for burn wound coverage.

Cells, Cultured↗