Direct current reduces accumulation of Evans Blue albumin in full-thickness burns.

BACKGROUND: Application of direct current (DC) to a burn wound limits extension of the zone-of-stasis and reduces wound tissue edema. OBJECTIVE: To study the effects of DC on extravasation of plasma proteins after burn by using Evans blue (EB) as a marker of plasma albumin. MATERIALS AND METHODS: Male Sprague-Dawley rats with 20% total body surface area full-thickness scalds (100 degrees C/10 sec) were used as the experimental model. Burn wounds were treated with plain nylon, silver-nylon, silver-nylon and 40 microA DC, or no dressing. EB (30 mg/kg) was injected immediately or at variably delayed postburn (PB) times and accompanied by DC application at various time intervals PB. Tissue content of Evans blue was assessed at different times after injection of the dye or infliction of burn injury. RESULTS: Evans blue albumin (EBA) concentration in untreated burn wounds (307.7 microg/g tissue) was nine times greater than in unburned skin (36.5 microg/g tissue) at 48 hours PB. When animals received a DC and EB injection immediately PB, DC treatment reduced EBA concentration by 60% at any time point PB. When EB was injected immediately PB, or at variably delayed times PB, accompanied by DC immediately PB, or at variably delayed times PB, DC reduced EBA accumulation at all examined times PB by more the 50% (p < 0.001). CONCLUSION: EBA and edema fluid accumulation in burn wound change in concert after injury and show similar response to DC treatment.

Optimized mesh expansion of composite skin grafts in rats treated with direct current.

BACKGROUND: The purpose of this study was to determine the optimum autoepidermal and allodermal expansion ratio of each component of a meshed composite skin graft (MCSG) that would lead to successful healing. METHODS: Male Sprague-Dawley rats were used as hosts of the MCSG and donors of autologous tissue. Male ACI rats were used as donors of allodermis. MCSGs with open meshed area (autoepidermal/allodermal) of 9:1/1.5:1, 9:1/3:1, 9:1/6:1, or 6:1/6:1 were applied to full-thickness skin defects and treated with a silver nylon dressing (SN) or SN with direct current (DC). Wound size, hair regrowth, and thickness of dermal layer were evaluated at 3 months. RESULTS: MCSGs of 9:1/1.5:1, 9:1/3:1, and 6:1/6:1 mesh ratios healed completely within 3 months with no difference in wound size between SN dressing groups or DC-treated groups. Application of DC reduced MCSG contraction and stimulated regrowth of hair. CONCLUSION: Fresh autoepidermis can be expanded 6:1 on a 6:1 allodermis or 9:1 on a 3:1 allodermis and achieve successful wound healing.

Direct current reduces plasma protein extravasation after partial-thickness burn injury in rats.

OBJECTIVE: To observe the effect of 40 microA direct current (DC) on plasma albumin extravasation after burn injury. DESIGN, MATERIALS, AND METHODS: Silver-nylon wound dressings were used as anodes (-) on anesthetized male Sprague-Dawley rats with 20% total body surface partial-thickness scald burns. Burned rats with no treatment, or treated with silver-nylon dressing without current, were used as controls. Quantitative analysis of fluorescein isothiocyanate (FITC)-albumin leakage and accumulation in the wound tissue was performed using confocal fluorescence microscopy. MEASUREMENTS AND MAIN RESULTS: In controls, the rate of albumin leakage was maximal at 1 hour postburn (PB) and then decreased, but remained higher than normal for 48 hours PB. The accumulation of FITC-albumin was maximal 4 to 6 hours PB and substantial for 48 hours. When DC was applied, leakage was reduced by 30 to 45% and approached normal control rates by 8 hours PB. FITC-albumin concentration peaked 4 hours PB, was 18 to 48% less then in burned control, and approached the level observed in unburned control by 18 hours PB. CONCLUSIONS: DC has a beneficial effect in reducing plasma protein extravasation after burn injury.

Direct current reduces wound edema after full-thickness burn injury in rats.

OBJECTIVE: To observe the effect of 4 and 40 microA direct current (DC) on edema formation after burn injury in rats. DESIGN, MATERIALS, AND METHODS: Silver-nylon wound dressings were used as either anodes (-) or cathodes (+) on 20% total body surface area full-thickness scalds in anesthetized male Sprague-Dawley rats. Untreated burned rats and rats treated with silver-nylon dressings without current were used as controls. MEASUREMENTS AND MAIN RESULTS: Immediately applied, continuous DC reduced burn edema by 17 to 48% at different times up to 48 hours postburn (p < 0.001). Neither reversal of electrode polarity nor change in current density had any significant effect on the results of treatment. Starting treatment during the first 8 hours postburn produced the least edema accumulation, but the reduction was significant even when DC was applied 36 hours afterburn. If started immediately after injury, treatment had to be continued a minimum of 8 hours to be most effective. CONCLUSIONS: Direct electric current has a beneficial effect in reducing wound edema after burn injury.

Enhanced survival of autoepidermal-allodermal composite grafts in allosensitized animals by use of silver-nylon dressings and direct current.

OBJECTIVE: Observe the effect of silver-nylon (SN) dressing and direct electric current on healing of meshed autoepidermal/allodermal composite skin grafts (MCSGs) in allosensitized rats. MATERIALS AND METHODS: MCSGs were placed on experimental animals 28 to 30 days after placement of sensitizing allografts. MCSGs and control allografts were covered with either Vaseline gauze (VG) or SN; direct current, 40 microA, was applied for 5 days to some of the SN-dressed wounds (SNDCs). MEASUREMENTS AND MAIN RESULTS: Second set rejection of MCSG was not observed. SN- and SNDC-treated grafts showed expansion of the meshed autoepidermis with complete epithelialization within 3 weeks. VG-covered wounds developed areas of open granulation and were not completely epithelialized at 3 months. Both SN and SNDC reduced wound contraction when compared to VG (SN versus VG p < 0.02, SNDC versus VG p < 0.008). MCSG was found to be of low alloantigenicity in that it did not induce second set rejection of subsequent skin allograft. CONCLUSIONS: SN dressings enhanced survival of meshed composite skin grafts.

The use of flow cytometry for the investigation of cell death.

Flow cytometry is more and more widely used for investigations of cell death, predominantly in the study of DNA degradation in cells dying by apoptosis. There are different interpretations of changes observed in DNA histograms of these cells. We describe an approach based on extraction of chromatin degradation products from fixed cells and subsequent staining with DNA specific dyes. Apoptotic cells containing fragmented DNA are observed in < 2C DNA region of DNA histograms. DNA histograms of irradiated thymocytes dying in vitro and stained without extraction of fragmented DNA do not differ from control. Under the same staining conditions DNA histograms of lymphocytes dying in thymus of irradiated animals reveal fluorescent material in < 2C DNA region, most likely due to formation of apoptotic bodies (cell fragments, some of them contain fragments of nuclei). Similar changes are observed in thymocytes dying upon glucocorticoid treatment. Our present results and other data indicate that reduced amount of DNA in dying cells is the main reason for changes of DNA histograms. Examples of application of the method described for the investigations of cell death modifiers are presented.

[The combined action of gamma irradiation and hyperthermia on the structure of the cell population in the Chinese hamster]. / Kombinirovannoe deistvie gamma-oblucheniia i gipertermii na strukturu populiatsii kletok kitaiskogo khomiachka.

A change in the structure of FAF-28 Chinese hamster cell population occurred during 24 h following gamma-irradiation or hyperthermia heating, or the effect of both factors was studied by flow cytofluorometry. With radiation delivered immediately after heating the distribution of cells among cycle phases was nearly the same as with hyperthermia alone: the share of cells at the S-phase was invariable during the first 4-6 h, then it slowly diminished; at G1 it slowly decreased and at G2 increased. When irradiation preceded heating the pattern of cell redistribution during the first hours was the same as that with radiation alone: the "wave" of transition from G1 to S phase was the same, but shorter in amplitude and longer in time; then cells were accumulated at G2+M and remained there for 24 h. Thus, of the two factors applied, the first was the major one in changing the cell population structure during the first hours after treatment. In 24 h the result was the same, that is, the considerable accumulation of cells at G2+M.

[The programmed cell death of murine BW5147 thymoma under the action of dexamethasone]. / Programmiruemaia gibel' kletok timomy myshi BW5147 pod deistviem deksametazona.

By flow cytometry, imitation modelling and biochemical analysis, the mode and kinetics of dexamethasone-treated T-lymphoma cell death were studied. The hormone was shown to induce delays in pre- and postsynthetic phases of the cell cycle and the death of part of cells. A short exposure to dexamethasone reveals its cytostatic rather than cytolytic effect. Following G2/M delay and cytokinesis, part of cells dies. A reduced serum concentration (2%) causes shorter delays in the cell cycle and a more rapid cell death. Dexamethasone stimulates apoptosis which is indicated by internucleosomal DNA fragmentation, and by a coincidence in time of the processes of DNA degradation and increase in the other membrane permeability. These results are discussed in relation to the cell death and proliferation.

Apoptosis of murine BW 5147 thymoma cells induced by dexamethasone and gamma-irradiation.

The mode and the kinetics of the death of T-thymoma cells upon dexamethasone treatment and gamma-irradiation (10Gy) have been studied using flow cytometry and biochemical analysis. It has been shown that the hormone and gamma-irradiation induce cell death by apoptosis. In both cases the cells are initially blocked in G2/M and die only after overcoming the blockage and cytokinesis. A short exposure to dexamethasone results in a cytostatic effect, whereas a cytotoxic effect is absent. Reducing serum concentration to 2% causes more rapid death both following gamma-irradiation and dexamethasone. These results are discussed in relation to cell death and proliferation.

[D2O inhibition of interphase thymocyte death]. / Ingibirovanie D2O interfaznoi gibeli timotsitov.

Using the method of flow cytometry and biochemical analysis it was shown that D2O, an agent that stabilizes microtubules, prevented the internucleosome fragmentation of DNA in thymocytes exposed to gamma radiation and dexamethasone in vitro. It was also found that D2O is ineffective with respect to Ca2+/Mg2(+)-dependent nuclease. The transfer of irradiated cells from a medium containing 90% of D2O to a normal one caused rapid DNA degradation; the fragmentation process ceased with the irradiated cells being transferred from H2O to heavy water. The results obtained permit us to assume that the disturbance of microtubules is not a trigger mechanism of DNA degradation by apoptosis, but is some intermediate stage of cell death preceding the chromatin fragmentation proper.

[DNA replication in mammalian cells after the action of physical, chemical or biological factors. VI. The recovery of DNA synthesis in a culture of irradiated cells]. / Replikatsiia DNK v kletkakh mlekopitaiushchikh posle deistviia faktorov fizicheskoi, khimicheskoi ili biologicheskoi prirody. VI. Vosstanovlenie sinteza DNK v kul'ture obluchennykh kletok.

The kinetics of DNA synthesis restoration in cultured HeLa cells and in L-929 mouse fibroblasts irradiated by gamma-rays of 60Co with a dose of 10 Gy was studied. Early after irradiation the rate of DNA synthesis in HeLa cells measured with 3H-thymidine incorporation was seen to decrease. Two hours later the incorporation starts to increase to reach the control level 4 hours after irradiation and then becomes even higher than this level. The distribution of cells among phases of the cell cycle measured with flow cytometry undergoes changes. 4-6 hours after irradiation part of S-phase cells increased contributing presumably to the elevating of 3H-thymidine incorporation observed at this time. The restoration of the incorporation was suppressed by inhibitors of protein and RNA synthesis--cycloheximide and actinomycin D. It is suggested that the processes of restoration of DNA synthesis in irradiated cells can be of inducible nature. In irradiated HeLa and L-929 cells the restoration of DNA synthesis is resistant to novobiocin, an inhibitor of DNA replication.

[Reproductive death of the apoptosis type in irradiated cells of the BW5147 thymoma]. / Reproduktivnaia gibel' obluchennykh kletok timomy BW5147 po tipu apoptoza.

The method of flow cytofluorometry and biochemical analysis were used to study the pattern and kinetics of the postirradiation death of proliferating BW5147 lymphoid cells. Irradiation with a dose of 10 Gy was shown to induce thymoma cell death by apoptosis. Radiation-induced synchronous transfer of part of cells from G1 to S-stage and blocking of all cells at G2/M stages of the cell cycle preceded the cell death. Decreasing of the growth factor content in a medium through its depletion or cultivation in conditions of low serum content accelerated cell death. A possible relationship between cell death and proliferation is discussed.

[Analysis of distribution of S. cerevisiae cells for protein content using cell cycle models]. / Analiz razpredelenii kletok S. cerevisiae po soderzhaniiu belka pri pomoshchi modelei kletochnogo tsikla.

Histograms of cell distributions according to protein content obtained by means of flow cytofluorometry characterize the physiological state of the population as a whole and permit to calculate the velocity of protein accumulation in the cell in the course of the cell cycle. Dependence of population heterogeneity on culturing conditions is considered. Mathematical analysis of histograms of continuous cultures of S. cerevisiae is carried out at dilution rates 0.4 hours-1 and 0.05 hours-1. Calculations are carried out on condition that the protein content in the cell rises a) exponentially and b) linearly in the course of the cell cycle. At low growth rate (0.05 hours-1) the distribution is bimodal and therefore it is highly informative. The assumption concerning linear accumulation of the protein allows good approximation of the experimental distributions by the theoretical ones.

[General similarities and differences in the postradiation death of various animal species and of man]. / Obshchie zakonomernosti i razlichiia postradiatsionnoi gibeli limfotsitov raznykh vidov zhivotnykh i cheloveka.

It was found in various animal species and man that an ordered internucleosome fragmentation of DNA is characteristic of lymphoid cells dying in the interphase. Both in vivo and in vitro, the postirradiation DNA degradation in thymocytes of rodents and piglets preceded the increase in the permeability of their plasma membrane. The in vivo kinetics of death of lymphoid cells from the thymus and spleen is similar in rodents and piglets. Rat thymocytes died in vitro earlier than thymocytes of piglets, calves and man which was evidently associated with a worse adaptive capacity of the latter to cultivation conditions.

[The patterns of colchicine-induced death of lymphoid cells]. / Zakonomernosti indutsirovannoi kolkhitsinom gibeli limfoidnykh kletok.

It is shown that colchicine injection at doses higher than 1 mg/kg of animal weight induces cell death in thymus, spleen, bone marrow and intestine mucosa. The cell death is accompanied by a regular internucleosomal cleavage of nuclear DNA and by the elimination of the formed fragments from cells. Both the processes begin after a 1.5 hour lag-period and proceed before the outer membrane permeability for supravital dyes increases. DNA degradation is prevented by the inhibitor of protein synthesis cycloheximide. Cytochalasin B does not induce chromatin degradation or cell death and has no effect on radiation death of lymphocytes. A possible role of microtubule destruction as a switch-on mechanism of DNA degradation and cell death is discussed.

[Interaction of the fluorescent probe diS-C3-(5) with lecithin-cholesterol membranes]. / Vzaimodeistvie fluorestsentnogo zonda diS-C3-(5) s letsitin-kholesterinovymi membranami.

Behaviour of fluorescent carbocyanine probe disS-C3(5) in the egg lecithin-cholesterol membrane suspension was studied in relation to the lecithin/cholesterol ratio. The partition coefficient of the probe between aqueous and lipid phases decreases unlinearly with increase of cholesterol molar part in a bilayer. This parameter over molar part units was estimated to be (2.4 +/- 0.1) X 10(6) for egg lecithin membranes and (1.8 +/- 0.2) X 10(6) for 10 mol% cholesterol, (1.2 +/- 0.1) X 10(6) for 20, (0.8 +/- 0.1) X 10(6) for 30, and (0.48 +/- 0.02) X 10(6) for 50 mol% cholesterol. It is suggested that the probe partition coefficient value consists of two components: one caused by pure lecithin bilayer regions and another by local lecithin concentration fluctuations in the mixed lecithin-cholesterol regions.