Arthur K. Balin

Oxidative influence on development and differentiation: an overview of a free radical theory of development.

Metabolic gradients exist in developing organisms and are believed to influence development. It has been postulated that the effects of these gradients on development result from differential oxygen supplies to tissues. Oxygen has been found to influence the course of development. Cells and tissues in various stages of differentiation exhibit discrete changes in their antioxidant defenses and in parameters of oxidation. Metabolically generated oxidants have been implicated as one factor that directs the initiation of certain developmental events. Also implicated as factors that modulate developmental processes are the cellular distribution of ions and the cytoskeleton both of which can be influenced by oxidants. The interaction of oxidants with ion balance and cytoskeleton is discussed.

Grafting of skin ulcers with cultured autologous epidermal cells

We treated five adult individuals with six full-thickness chronic ulcerations in the skin caused by venous insufficiency, sickle cell anemia, or surgical wounds. Each patient received applications to the ulcerations of sheets of autologous epidermal cells grown in culture. All patients experienced relief of pain after grafting. Four of the six ulcers healed completely in 21 to 35 days, and three of the four remained healed for up to 2 years. One ulceration recurred within 2 months. Our experience suggests that cultured autologous epidermal grafts can provide continuous covering, relief from pain, and rapid healing of chronic debilitating ulcerations of the skin.

Superoxide dismutase induces differentiation in microplasmodia of the slime mold Physarum polycephalum

Evidence is presented that supports a role for the enzyme superoxide dismutase (SOD) in the differentiation of the slime mold, Physarum polycephalum. SOD activity increases 46-fold during differentiation. A strain of Physarum that does not differentiate exhibits no change in SOD activity. Addition of SOD, via liposomes, to the nondifferentiating strain induces differentiation; this effect is enhanced by an inhibitor of glutathione synthesis. Other antioxidants selected for study failed to induce differentiation. Conversely, oxidative treatments including introduction of D-amino acid oxidase, via liposomes, induced differentiation. Cellular oxidation is the probable cause of the SOD effect.

Effects of oxygen on the antioxidant responses of normal and transformed cells

Basal antioxidant defense levels are often aberrant in tumor cells; however, less attention has been given to differences in the way that normal and transformed cells respond to changes in oxidative stress. This study evaluated differences in the responses of various normal and transformed cell lines to different oxygen tensions. Exposure to hyperoxia generally failed to induce either the activity of GSH peroxidase (GPx) or the manganese-containing form of superoxide dismutase (MnSOD) after 48 h, although at 605 mm Hg oxygen, small inductions of MnSOD activity were observed in adult lung fibroblasts and amelanotic melanoma. Exposure to 605 mm Hg O2 for 48 h was inhibitory to GPx activity. MnSOD activity was strongly induced in virally transformed WI-38 cells by treatment with the herbicide paraquat or inhibition of GSH synthesis with BSO. In normal cells GSH concentration was proportional to ambient oxygen tension. Tumor cells exhibited greater GSH concentrations at low oxygen tensions than normal cells but were unable to increase GSH in response to elevation of oxygen tension. These results reveal differences in tumor and normal cell responses to changes in ambient oxygen tension and show that MnSOD activity is inducible when an appropriate stimulus is applied.

Effects of ambient oxygen concentration on the growth and antioxidant defenses of of human cell cultures established from fetal and postnatal skin.

Oxygen toxicity is believed to arise from changes in the rates at which cells generate reactive oxygen species (ROS). Sensitivity to hyperoxia has been postulated to depend on levels of antioxidant defense. Human cells obtained from fetal tissues have lower antioxidant defenses than those obtained from adult tissue. The present study was performed to determine whether the differences in fetal and adult antioxidant defense levels modulated their responses to changes in the ambient oxygen concentration. Our results demonstrate that oxygen modulates the proliferation of human fetal and adult skin fibroblasts in a similar fashion. In general, skin fibroblasts grew better at approximately 31 mm Hg, regardless of donor age. Manganese superoxide dismutase, catalase, and glutathione peroxidase activities were lower in fetal cells than in adult fibroblasts. Copper/zinc superoxide dismutase and glucose-6-phosphate dehydrogenase were similar in fetal and postnatal tissues and were unaltered appreciably by hyperoxic exposure. Glutathione concentration increased at higher oxygen tensions; however, the increase was much greater in fetal cells than in cultures derived from adult skin. These observations demonstrate that the capacity of fetal and adult cells to cope with oxidative stress, while similar, result from distinct mechanisms.

Normal replicative lifespan of Alzheimer skin fibroblasts.

Cultured skin fibroblasts from four patients with Alzheimers disease had life spans comparable to those of six age-sex matched controls, whether measured by passages to phase-out, cumulative population doublings to phase out, or percentage of nuclei incorporating [3H]thymidine (Cristofalo index). These results provide direct experimental evidence that Alzheimers disease is not simply a form of accelerated aging. They suggest that the abnormalities, described by several groups, in Alzheimer fibroblasts reflect the disease rather than the physiological age of the donor, making the cultured cell a valid tool for studying the cellular pathophysiology of this disorder. Together with other data, these observations raise the possibility that some forms of Alzheimers disease may represent inborn error(s) of metabolism of late clinical onset.

Effects of static magnetic fields on the growth of various types of human cells

The effects of a static magnetic field (SMF) on the proliferation of various types of human cells were determined. All cultures were maintained at 37 °C throughout the experiment. SMF was generated by placing two magnets oppositely oriented on either side of a T25 flask. The flux density in the flask ranged from 35 to 120 mT. Growth curves were constructed by plotting cell number at 18 h and 4, 7, 11, and 14 days after seeding, with the 18-h point being a measure of attachment efficiency. Exposure to SMF significantly decreased initial attachment of fibroblasts and decreased subsequent growth compared to sham-exposed control. Significant effects were observed in both fetal lung (WI-38) and adult skin fibroblasts, but they were generally larger in the fetal lung fibroblast line. SMF did not affect attachment of human melanoma cells, but inhibited their growth by 20% on day 7. SMF produced no effects in a human adult stem cell line. Oxidant production increased 37% in WI-38 cells exposed to SMF (230-250 mT) during the first 18 h after seeding, when cell attachment occurs. Conversely, no elevation in oxidant levels was observed after a prolonged 5-day exposure. These results indicate that exposure to SMF has significant biological effects in some, but not all types of human cells.

OXYGEN MODULATES THE GROWTH OF SKIN FIBROBLASTS

SummaryElevated oxygen tensions are inhibitory to the growth of skin fibroblasts. Skin fibroblasts grow better at oxygen tensions below 137 mm Hg regardless of seeding density. A wide range of oxygen tensions, including those in the physiological range, strongly modulate the growth of human skin fibroblasts. There were no significant differences between the responses of fetal and postnatal cell lines to changes in ambient oxygen tension. In all cases, higher oxygen tensions significantly impeded cell growth. Seeding cells at 104 cells/cm2 afforded some protection from the deleterious effects of hyperoxia. Oxygen tensions exceeding the amount present in ambient room air also impeded cell growth at this higher seeding density, but the effect did, not become significant until the oxygen partial pressure reached 241 mm Hg. At lower oxygen tensions, cells seeded at 103 cells/cm2 grew more rapidly than did cells seeded at 104 cells/cm2. These findings may have implications for the treatment of poorly healing wounds with hyperbaric oxygen.

Cannulation and Injection of the Muscles of Facial Expression: A Cadaver Study

BACKGROUND Facial fat grafting typically places autologous fat in the subcutaneous plane directly under the surfaces that are deemed to need volume augmentation. Because this plane is relatively avascular, there can be variable loss of the fat graft. Placing the fat into a rich muscular vascular plexus may increase graft retention. OBJECTIVES To explore the feasibility of engrafting the muscles of facial expression in the plane and direction of their fibers using a percutaneous approach. MATERIALS AND METHODS Three muscles—corrugator, zygomaticus major, and depressor anguli oris—were chosen for this study. More than 50 thawed frozen cadaver heads were studied. A blunt‐tipped cannula, designed to find and follow the muscle within its enveloping fascia, was introduced through an access port created through the skin using an awl. One to 2 mL of blue gel was injected per muscle from a 3‐mL syringe. Immediately after injection, the muscle was dissected to determine placement of the blue gel. RESULTS Accurately targeting the muscles of facial expression for autologous fat grafting using a percutaneous approach is feasible. CONCLUSION With practice, the muscles of facial expression can be reliably cannulated percutaneously, permitting restoration of volumetric losses. Dr. Amar has a patent on the cannulas used in this study.

Human Fibroblast Antioxidant Defense Response to Alteration in Oxygen Tension

Our previous studies have revealed that partial pressures of oxygen (PO2) ranging from 6 through 720 mm Hg modulate growth of human diploid fibroblasts. We have demonstrated that even physiological oxygen concentrations (PO2 6 – 134 mm Hg) modulate the proliferative life span of these cells. Furthermore, proliferatively aged cells that exhibit a decreased growth rate are more sensitive to oxidative stress than are actively growing cells at lower population doubling levels. Inhibition of cell growth by partial pressures of oxygen above PO2 300 mm Hg and the limited proliferative capacity of cells in culture under physiological oxygen concentrations are probably distinct phenomena resulting from different mechanisms. Our studies indicate that fibroblasts contain multiple oxygen-sensitive sites which exhibit differential sensitivity to oxygen; however, total cellular metabolism is not generally inhibited by hyperoxic exposure. Thus, oxygen-induced inhibition of cell growth may result from a relatively selective process.1