Kathleen M. Miller

Biomaterial biocompatibility and the macrophage.

The biocompatibility of biomaterials at implant sites is controlled by the tissue/material interaction. A major cell in the tissue reaction is the macrophage. A summary is presented on macrophage mediation of cellular and humoral regulatory pathways in inflammatory and immune responses.

In vitro and in vivo interactions of cells with biomaterials

The biocompatibility of materials at an implant site involves a complex interaction of cells and tissues with the biomaterial. This cell-cell and cell-polymer interaction evokes the release of mediators such as chemotactic and growth factors that elicit and sustain inflammatory responses at the implant site. In this review, we summarize the interaction of cells with biomaterials in vitro and in vivo.

O6 alkylguanine-DNA alkyltransferase activity in human myeloid cells.

The association between alkylating agent exposure and acute nonlymphocytic leukemia in humans indicates that myeloid cells may be particularly susceptible to mutagenic damage. Alkylating agent mutagenesis is frequently mediated through formation and persistence of a particular DNA base adduct, O6alkylguanine, which preferentially mispairs with thymine rather than cytosine, leading to point mutations. O6alkylguanine is repaired by O6alkylguanine-DNA alkyltransferase (alkyltransferase), a protein that removes the adduct, leaving an intact guanine base in DNA. We measured alkyltransferase activity in myeloid precursors and compared it with levels in other cells and tissues. In peripheral blood granulocytes, monocytes, T lymphocytes, and B lymphocytes, there was an eightfold range of activity between individuals but only a twofold range in the mean activity between cell types. Normal donors maintained stable levels of alkyltransferase activity over time. In bone marrow T lymphocytes and myeloid precursors, there was an eightfold range of alkyltransferase activity between donors. Alkyltransferase activity in the two cell types was closely correlated in individual donors, r = 0.69, P less than 0.005, but was significantly higher in the T lymphocytes than the myeloid precursors, P less than 0.05. Liver contained the highest levels of alkyltransferase of all tissues tested. By comparison, small intestine contained 34%, colon 14%, T lymphocytes 11%, brain 11%, and myeloid precursors 6.6% of the activity found in liver. Thus, human myeloid precursors have low levels of O6alkylguanine-DNA alkyltransferase compared with other tissues. Low levels of this DNA repair protein may increase the susceptibility of myeloid precursors to malignant transformation after exposure to certain alkylating agents.

Characterization of biomedical polymer-adherent macrophages: interleukin 1 generation and scanning electron microscopy studies

Macrophage activation following attachment to biomedical polymers was studied using two systems of analysis. Supernatants generated by human peripheral blood monocytes cultured on the surface of several different biomedical polymers were evaluated for the presence of the secreted regulatory protein interleukin 1 (IL1). In addition, each cell-polymer culture surface was subjected to scanning electron microscopy for gross morphological evaluation. Results indicate that, although all materials were efficient in the attachment and activation of cells, the panel of polymers showed a differential capacity in attachment and activation of monocytes. Dacron and polyethylene surfaces had a greater density of cells showing morphology indicative of activation, corresponding to elevated levels of IL1 in these cultures. Biomer and polydimethylsiloxane surfaces showed fewer activated cells and had lower IL1 levels in culture. Expanded polytetrafluorethylene resulted in intermediate levels of IL1 and attached cells showing activated morphology.

Normal lymphocyte responses to mitogens in term and premature neonates following normal and abnormal intrauterine growth

Cord blood lymphocyte responses to a panel of four mitogens were studied in 242 neonates using a whole blood technique. The patient population was divided into five gestational-age groups: 20-27.9, 28-32.9, 33-37.9, 38-41.9, and 42-44 weeks. Neonatal lymphocytes undergo a continuous reduction in proliferative responsiveness to the polyclonal ligands phytohemagglutinin (PHA) and concanavalin A (Con A) as gestation progresses from 20 to 44 weeks postconception. This is consistent with their change in unstimulated in vitro blastogenesis which when measured over the same developmental period is greatest in more immature neonates. Neonatal lymphocyte proliferative responsiveness to pokeweed mitogen (PWM) and staphylococcus protein A (SpA), however, was unrelated to gestational age. The influence of intrauterine nutritional deprivation on lymphocyte proliferative responses was studied in clinically uninfected newborns and compared to gestational age-matched controls with a normal nutritional status. Intrauterine nutritional deprivation was not associated with a decrease in mitogen-induced lymphocyte proliferation. Further, we explored the influence of several perinatal clinical settings commonly associated with fetal distress on cord blood lymphocyte responses to mitogens. Although perinatal stress in the form of low Apgar scores, meconium-stained amniotic fluid, and prolonged rupture of the amniotic membranes was not related to differences in mitogen induced lymphocyte proliferation, mode of delivery was. Cesarean section delivery as compared to vaginal delivery was associated with a significantly greater PHA-, Con A-, and SpA-induced neonatal lymphocyte response. Several alternative explanations for this finding are explored. Lastly, the purified protein thymosin fraction 5 was not associated with alteration in either neonatal or adult mitogen-induced lymphocyte proliferation.

Perinatal influences on in vitro B lymphocyte differentiation in human neonates.

ABSTRACT: In vitro differentiation of B lymphocytes present in cord blood mononuclear cell preparations into immunoglobulin secreting cells was studied in 126 neonates with gestational ages (GA) ranging from 20 to 44 wk. Eight infants had a GA less than 27.9 wk, 24 had GA 28–32.9 wk, 30 had GA 33–37.9 wk, 51 had GA 38–41.9 wk, and 13 had GA above 42 wk. B cell differentiation in response to pokeweed mitogen plus hydrocortisone was assessed using a plaque forming cell assay. All neonates had a measurable plaque-forming cell response in this assay. An increased plaque-forming cell response was observed in some neonates in all gestational age groups. The magnitude of in vitro neonatal B cell differentiation underwent a continuous and significant (p < 0.002) reduction as gestational age increased. The influence of intrauterine growth retardation on in vitro B lymphocyte differentiation was studied and compared to gestational age-matched controls with a normal intrauterine growth. Intrauterine growth retardation was not associated with changes in B cell responsiveness. An analysis of perinatal factors revealed that cesarean section, and low 1-min Apgar scores were factors that predisposed cord blood cells to be triggered in vitro to produce increased numbers of plaque-forming cells.

Selected Aspects of Cell and Molecular Biology of In Vivo Biocompatibility

The biocompatibility of an implanted material or prosthetic device is a dynamic and two-way process that involves the time dependent effects of the host on the material and the material on the host. The implantation of any synthetic material initiates a wound healing mechanism that is characterized by the inflammatory response. However, the inflammatory response itself involves complex and highly regulated interactions between specific cells and various molecular mediators. An understanding of these interactions which occur following biomaterial implantation has been hindered by the difficulty in quantifying the cellular and biological events.

1017 INFLUENCE OF LABOR ON CORD BLOOD LYMPHOCYTE (1ym) POPULATIONS

We have previously demonstrated that cord blood lym proliferative responses(J Clin Immuno/Immunopath, 30, 1984) and levels of pokeweed induced antibody secreting cells (Pediatr Res, in press) are greater among neonates delivered by Cesarean section(CS) than among those delivered vaginally. These increases were noted to be related to the absence of labor prior to CS. To determine if the presence of labor prior to delivery influenced the numbers and/or proportions of cord blood(CB) T-lym populations, these cells were identified in 46 term neonates (17 delivered vaginally, 29 CS). Mononuclear cells were isolated on a ficollhypaque gradient. T-lym were identified with the monoclonal antibodies OKT3(all T-lym), OKT4(helper-inducer lymphocytes) and OKT8 (suppressor/cytotoxic T cells) using flow cytometry after indirect immunoflucrescent labeling.*p values: labor vs no labor <.002, vaginal vs no labor <.0013 These data suggest that labor influences the proportions of CB T-lym subsets without changing the total number of lym. Changes in T-lym subpopulations could explain differences in T and B lym responses in newborns delivered by CS.

NEONATAL B CELL DIFFERENTIATION (RIFF)

Perinatal factors influencing diff of cord blood B lymphocytes into immunoglobulin secreting plasma cells was studied in 126 neonates with gestational ages(GA) from 20-44 wks. A plaque forming cell assay measured background B cell diff and diff in response to pokeweed mitogen(PWM)plus hydrocortisone(HC). 8 infants had a GA less than 27.9 wks, 24 had GAs 28-32.9 wks, 30 had GAs 33-37.9 wks, 51 had GAs 38-41.9 wks and 13 had GAs above 42wks. The mean±SD GA was 36±5wks and B.Wt. 2400±980gm. The mean±SD background plaque formation was 98±253 and in response to PWM+HC was 7505±11874/106 cord blood mononuclear cells. B cell diff was observed in some neonates in all GA groups. The magnitude of in vitro neonatal B cell diff undergoes a continuous and significant(p<.002)reduction as GA increases. The influence of intrauterine nutritional deprivation(IUGR)on B lymphocyte diff was studied, and compared to GA matched controls with a normal intrauterine nutritional status. IUGR was not associated with a decrease in B cell responsiveness. Cesarean section and low one minute Apgar scores were associated with a significantly(p<.05)increased B lymphocyte diff. Although prolonged rupture of maternal amniotic membranes(PROM)was not singularly related to a significant increase in B cell diff, the combination of PROM and low Apgar scores(stress)was associated with a more significant increase(p<.003)than either factor alone. Thus, short term neonatal stress such as the combination of PROM and low Apgar scores has as much influence on neonatal B cell function as GA.

MODE OF DELIVERY AND NEONATAL LYMPHOCYTE PROLIFERATION|[lpar]|LP|[rpar]|

Neonatal mitogen(M)induced lymphocyte proliferation reflects the hosts ability to develop a cellular immune reaction. Since LP and lymphokine release usually occur simultaneously and lymphokines enhance phagocytosis, LP suggests resistance to infection. To determine the influence of delivery on LP, we measured LP in vitro in 175 vaginally(vag) and 65 Cesarean section(CS)delivered neonates. LP/105 cord blood mononuclear cells was measured with a whole blood assay measuring tritiated thymidine incorporation(cpm). LP was significantly(p<.02)greater in neonates delivered by CS than in those delivered vag, and was more significantly increased in neonates delivered to mothers with no labor (cervical dilatation and/or effacement with uterine contractions) preceding delivery. This difference was nor related to maternal anesthesia or perinatal stress (Apgar scores 61).Within the group of neonates delivered by CS, prior labor significantly(p<.04)decreased LP. Thus, the presence of labor prior to delivery significantly influences neonatal LP and may influence neonatal host defense.a=mean±SD; b=mean±SD × 103cpm/105 cord blood mononuclear cells