E.S. Lindenbaum

Enhancing the survival of aspirated human fat injected into nude mice

Injection of aspirated fat is now the most commonly used technique for the filling of depressed areas. Partial absorption of the injected fat is the main limitation of this procedure. Cariel T.M. is an enriched serum-free cell culture medium, its ability to enhance the survival of human aspirated fat grafts was investigated in the nude mouse model. A volume of 0.75-cc Cariel preprocessed fat was injected under the scalp skin of 16 nude mice in the experimental group, and the same volume of saline preprocessed fat was injected to 15 control group of mice. Significant maintenance of the weight, 46 percent in the experimental group compared with 29 percent in the control group (p < 0.008), and the volume, 44 percent in the experimental group compared with 31 percent in the control group (p < 0.026), was observed, after 15 weeks, in this newly used model. It seems that addition of the nutrients enriched with anabolic hormones enabled the survival and take of more adipose cells in the graft.

Static-electric field induction by a silicone cushion for the treatment of hypertrophic and keloid scars

&NA; Silicone gel and silicone occlusive sheeting are widely used at present for the treatment of hypertrophic and keloid scars, without any scientific explanation as to their mode of action. In a recent paper the possibility was raised that static electricity generated by friction‐activated silicone sheeting could be the reason for this effect, and that it can, with time, cause involution of hypertrophic and keloid scars. The objective of this study was to test this hypothesis and to observe whether a continuous and also an increased negatively charged static‐electric field will shorten the treatment period. A device to implement these requirements gradually evolved over a 5‐year period. A number of prototypes were tested until the final product was attained. Some of the patients in this study were treated initially with a silicone sponge inserted in the cushion. Later this version was changed to the final design described herein. A silicone cushion was developed with the purpose of increasing a negative static‐electric charge to accelerate the regression process. The cushion is custom‐made using a silicone occlusive sheeting envelope of 0.75‐mm thickness, which does not deteriorate with use, and is partially filled with high viscosity silicone oil. Its edges are sealed, and its size is designed to extend a little beyond the scarred area. Static electricity readings, generated by activating the cushion by pumping action with the fingers, stretching or deforming the cushion, are invariably much higher when compared with those obtained with silicone occlusive sheeting and silicone gel sheeting. The interaction between the negatively charged ions of the cushion and the ionic charges of the tissue fluids may be the critical factor in achieving hypertrophic and keloid scars involution. Of the 30 patients enrolled in the study, 3 patients dropped out. Treatment with the silicone cushions yielded 63.3 percent cessation of itching and burning followed by pallor and flattening of the scar, some markedly so, over a few weeks to 6‐month period. An additional 26.6 percent had their scars resolved in up to 12 months of treatment. Good contact of the cushion over the scar has been shown to be important in this clinical trial, and much creativity is needed for making elastic strap bindings that ensure this contact. The clinical trials extended over a 12‐month period. Ten patients (33.3 percent) who had recalcitrant scars with little response to the use of the silicone cushion were given intralesional corticosteroid injections, in addition to the continued use of the cushion, resulting in a fairly rapid resolution of these scars over a period of months to a year. (Plast. Reconstr. Surg. 101: 1173, 1998.)

The two- and three-dimensional structure of the microcirculation of the chick chorioallantoic membrane.

The chick chorioallantoic membrane (CAM) is a common model for studying biological processes, but descriptions of the CAM circulatory system and especially experimental preparations of the CAM in shell-less eggs are both scant and controversial. We studied the CAM structure and the three-dimensional spatial configuration of the CAM vessels using five methods: in vivo stereoscopic observations, whole-mount preparations, histological sections, corrosion cast microinjection techniques, and the reconstruction of a three-dimensional wax model. Our findings show that the CAM consists of a superficial two-dimensional layer composed of a network of a very dense capillary mesh, floating over and enclosing a deeper three-dimensional space in which medium and larger free-floating vessels are seen to supply and drain the superficial layer. It is interesting to note that no tips or sprouts of blood vessels were observed during the development of the CAM vessels. In a shell-less egg preparation, the capillaries were found in the mesoderm layer of the CAM and not in or superficial to the ectoderm as is the case in the CAM of the intact egg which adheres to the shell membrane.

Serotonin and the ovarian hyperstimulation syndrome

Ovarian hyperstimulation syndrome was induced in rabbit by administration of human menopausal gonadotropin (HMG) and human chorionic gonadotropin (HCG). In an attempt to establish whether serotonin plays a part in the induction of this syndrome, the hyperstimulated rabbits were divided into two groups and were administered known anti-serotonin drugs, cyproheptadine and methysergide, respectively. The group treated with cyproheptadine, a non-specific serotonin antagonist, exhibited significant acceleration in the regression of the syndrome. Methysergide, a specific serotonin antagonist, administered to the second group, neither prevented the occurrence of the syndrome nor accelerated its regression. The results of this work indicate that serotonin does not seem to be directly involved in the production of the ovarian hyperstimulation syndrome in rabbits.

Improved vitality of experimental random dorsal skin flaps in rats treated with enriched cell culture medium

A defined, serum-free cell culture medium supplemented with nonsteroidal anabolic hormones, insulin, thyroxin, and growth hormone was found to accelerate wound healing by stimulating vascularized granulation tissue formation, epithelialization, and angiogenesis. The aim of this work was to study the effect of cell culture medium on the survival rate of cephalically based random dorsal skin flaps in an animal model. A total of 77 Sprague-Dawley rats were randomized into five treatment groups: pharmacologic delay with cell culture medium, flap enhancement with cell culture medium, surgical delay, biological delay with saline, and control. Statistically significant differences in distal flap necrosis were found among all groups (p<0.003). The rats treated with cell culture medium before flap elevation showed a significant increase in flap viability: a survival rate of 83 percent, compared with the control group, which demonstrated a survival rate of only 58 percent (p<0.0001). The surgical delay and the groups treated with cell culture medium yielded similar results with no significant difference between them. This study indicates that preoperative injection of cell culture medium may play a role in decreasing skin flap necrosis.

Computerized morphometric quantitation of elastin and collagen in SMAS and facial skin and the possible role of fat cells in SMAS viscoelastic properties

Recently, the superficial musculoaponeurotic system (SMAS) was found to be a composite tissue comprising collagen, elastic fibers, and fat cells in an extracellular viscous matrix. Both SMAS and facial skin tissues exhibit viscoelastic properties, but SMAS tissue has delayed stress relaxation. As a consequence, SMAS is viewed as a firmer elastic foundation for the more viscous facial skin. In some patients, a slackening effect of SMAS tissue takes place over a period ranging from weeks to months after tightening. To determine the relative quantity of viscoelastic components and better understand their biomechanical behavior, a quantitative morphometric study of the elastic and collagen fibers in the SMAS and facial skin was conducted. Thirty-four SMAS preparations were taken from 17 patients during either primary face lift operations (12 women) or reoperative face lift procedures (4 women, 1 man), which were performed 4 to 9 months after the original surgery, to examine the elastin and collagen content. For comparison, preauricular skin was also gathered from these patients. The specimens were stained with Weigerts staining to identify elastin and collagen fibers. Using a computerized morphometric analysis, 100 fields of each SMAS and skin specimen were examined. According to our findings, the average percentage of elastin and collagen fibers in SMAS and facial skin was as follows: (1) the percentage of elastin fibers in the SMAS was 4.71 +/- 1.2 (standard error of mean +/- 0.0291); (2) the percentage of elastin fibers in the skin was 6.1 +/- 1.8 (standard error of mean +/- 0.0436); (3) The percentage of collagen fibers in the SMAS was 38.7 +/- 5.9 (standard error of mean +/- 0.1430); and (4) the percentage of collagen fibers in the skin was 48.47 +/- 6.96 (standard error of mean +/- 0.1688). A statistical significance of p < 0.0001 was demonstrated between the collagen and elastin groups. A different percentage of elastin and collagen fibers was found among the 17 patients and within each of them separately. Neither gender nor age differences were found regarding elastin and collagen fiber content. No statistical differences were demonstrated between specimen sources, i.e., whether the operations were primary or reoperative face lift procedures. Findings from previous studies indicate that the cheek has two viscoelastic layers, the skin and the SMAS. The proportional similarity in average percentages of elastin and collagen in SMAS and facial skin cannot explain the relatively delayed stress relaxation effect of the SMAS. Therefore, the fat cells that are found exclusively in the SMAS probably lend a certain degree of firmness to this layer and play a significant role in the long-term efficacy of SMAS surgery.

Prolactin inhibits hCG-stimulated steroidogenesis and cAMP accumulation, possibly by increasing phosphodiesterase activity, in rat granulosa cell cultures.

The effects of prolactin (PRL), alone and together with human chorionic gonadotropin (hCG), on steroidogenesis and cAMP accumulation in the preovulatory ovary were studied. Cultured granulosa cells obtained from large preovulatory follicles of pregnant mare serum gonadotropin (PMSG)-treated immature rats were used. The results indicated that PRL inhibited, in a dose-dependent manner, hCG-induced cAMP accumulation and 17 beta-estradiol (E2) secretion. When added to 0.4 IU/ml hCG (designated as 100% activity), 1, 10 and 100 ng/ml PRL decreased cAMP accumulation to 86, 64 and 59%, respectively, following 1 h incubation and to 87, 81 and 66% E2 secretion, respectively, following 48 h incubation. PRL alone failed to cause any significant change in cAMP or E2 concentrations. The inhibition of PRL was apparently not at the hCG receptor level, since a similar inhibitory effect was observed in prostaglandin E1 (PGE1)-induced cAMP accumulation. Nor was the inhibitory pathway of adenylate cyclase involved, since pertussis toxin--an inactivator of the Gi regulatory protein--failed to abolish the suppressive effect of PRL on hCG-induced cAMP accumulation. The phosphodiesterase (PDE) inhibitor, 3-isobutyl-1-methyl-xanthine, abolished the inhibitory effect of PRL on hCG- and PGE1-induced cAMP accumulation and on hCG-induced E2 secretion, indicating that PRL might be inhibiting cAMP accumulation and steroidogenesis in preovulatory granulosa cells by enhancement of PDE activity.

Novel hyperactive mitogen to endothelial cells: human decidual NKG5.

Langer N, Beach D, Lindenbaum ES. Novel hyperactive mitogen to endothelial cells: human decidual NKG5. AJRI 1999; 42:263–272

Isolation and Culture of Human Decidual Capillary Endothelial Cells in Serum-Free Medium Supplemented with Human Uterine Angiogenic Factor

Human decidual capillary endothelial (HDCE) cells, obtained from decidual fragments of legally induced first-trimester terminations of pregnancies, were cultured in a serum-free medium supplemented with human uterine angiogenic factor (HUAF). The method of isolating the cells from the decidual tissue is described. Identification of the decidual endothelial cells was based on light- and electron-microscopic observations as well as on antifactor VIII immunoperoxidase-staining technique. The HDCE cell culture in the serum-free medium lasted for 25 weeks through eight subcultures. The cells frozen in glycerin yielded 80% viability after thawing. Electron-microscopic observations of the monolayer demonstrated Weibel-Palade bodies, desmosomes and tight junctions. HUAF is mitogenic to HDCE at 10-100 ng/ml. The dependency of HDCE cells grown in culture on HUAF may explain in part the mechanism involved in the dynamic vascular expansion occurring during gestation.

Role of mitogenicity in pathogenicity of mycoplasmas for murine hosts

The mitogenicity and pathogenicity of Mycoplasma pulmonis were compared in two rat strains. Both the mitogenic and the pathologic effects induced by M. pulmonis membranes were more severe in Lewis rats than in Hooded rats, and were dependent on the mitogen doses used. It was concluded that the severity of lung lesions induced by M. pulmonis membranes correlated with the degree of mitogenic responses of the different rat strains to this organism. The roles of T- and B-cell mitogens in induction of pneumonia were studied in Hooded rats treated intranasally with either the T-cell mitogen concanavalin A or with M. neurolyticum membranes which stimulate the B-cell populations, or with both concanavalin A and M. neurolyticum. Results clearly showed that the individual B- and T-cell mitogens affected the lungs of treated animals. Nevertheless, the mitogenic co-stimulation of both B and T lymphocytes in rat lungs was necessary to obtain maximal development of interstitial lymphocytic pneumonia.