<p>The development of murine gonads commences in both males and females as indifferent appearing blastemata around the 10th day of embryogenesis. Within the following two days, the, gonadal blastemata became heavily populated with the primordial germ cells which migrate in from extragonadal sites.</p> <p>The process of gonadal sex differentiation is first evident in males where the appearance of sex cords marks testicular histogenesis. Ovarian development is largely marked by the fact that female gonads do not resemble testes and also by the early transformation of oogonia into oocytes.</p> <p>It has been proposed by a number of investigators that the absence of the primordial germ cells in the gonadal blastemata during the indifferent stage of gonadogenesis would result in the failure of sex cord differentiation to take place within the gonadal soma. Other investigators have proposed that gonadal sex differentiation is not dependent upon the presence of germ cells within the gonadal soma.</p> <p>To resolve this controversy mice carrying the alleles Steel/Steel Dickie (S1/S1ᵈ) were selected since the Steel mutation is known to severely affect the primordial germ cell line prior to its populating the gonadal ridges before sex cord differentiation. The experiment therefore is a natural one not requiring chemical or physical ablation of the germ cell line at the time it is first histochemically identifiable in sites far removed from the gonadal anlages.</p> <p>Primordial germ cells were identified histochemically by an azo dye technique in 27 day 9, 33 day 10, 33 day 11 and 21 day 12 embryos which were obtained from matings between mature WC/ReJ-S1/+ females and C57B1/6J-S1ᵈ/+ males. One fourth of the embryo zygous S1/S1ᵈ mutation. Failure to the germ cell population to increase after 9 days gestation occurred in 26% of the embryos. These embryos were classified as the mutant S1/S1ᵈ group. The primordial germ cells of the mutants, though few in number, were found to follow a normal migratory pattern to the gonadal ridges. Gonadogenesis was studied in 227 fetuses of 12-18 days gestation and in animals from birth to maturity. The genetic sex of fetuses was determined by the presence or absence of sex chromatin in amnion cells. Prior to day 14, genotypes were established as normal or mutant according to the germ cell population present in one gonad from each fetus. After day 14 the genotypes were determined from red blood cell samples, the S1/S1ᵈ fetuses displaying the macrocytic anemia characteristic of this mutation.</p> <p>The ratio of mutant males to mutant females was the normal 1:1, and the number of mutants identified was within the expected 25% frequency. Despite a paucity of germ cells, mutant gonads differentiated according to their genetic sex. Although mutant gonads are composed almost entirely of somatic tissue they grow at the same rate as the somatic component of normal gonads up to day 16 post coitus in males, and to the day of birth in females. The overall pattern of growth was similar to that seen in normals. The Mullerian and Wolffian ducts as well as the external genitalia also developed according to gonadal sex in the mutants.</p> <p>In those mutant gonads which contained few germ cells, they grew and differentiated in the same manner as did those of the normals up to birth. By maturity, no mutant germ cells were found to have differentiated beyond prophase of meiosis I.</p> <p>It is concluded that sexual differentiation and gonadogenesis can take place in the absence or at least in the near absence of germ cells. The Steel mutation appears to act in preventing the proliferation of the primordial germ cells and their capacity to complete meiosis I of gametogenesis. The mutation, however, does not affect the evolution of the germ cells from their source nor the capacity of the germ cells to migrate to the gonadal blastemata.</p> / Doctor of Philosophy (PhD)

<p>Denervation of skeletal muscle fibres is known to lead to a marked alteration of their normal characteristics. The mechanism for this is not clear, but there is great interest in the possibility that the changes are the result of the cutting off of neurotrophic factors which normally continuously act on the muscle fibres. To study this question, colchicine, a drug known to interfere with neurorial transport, was injected through the epineurium of one sciatic nerve in rats, in an attempt to prevent the postulated neurotrophic factors from reaching the extensor digitorum longus (EDL) muscle, without interfering with impulse activity in the nerve, or the consequent activity of the muscle. Following this procedure, ipsilateral EDl muscle fibres were found to exhibit extrajunctional sensitivity to acetylcholine ( ACh ), tetrodotoxin ( TTX ) resistant action potentials, and lowered membrane potentials ( RMP ); all characteristic changes observed in denervated skeletal muscle fibres. These fibres however, were shown not to be denervated, since they displayed miniature end-plate potentials (m.e.p.p.s) and neuromuscular transmission was normal.</p> <p>Unexpectedly, the contralateral EDL muscle fibres which were examined as a routine control, also displayed the features of "denervation" following an injection of colchicine into the ipsilateral sciatic nerve, and were virtually indistinguishable from the ipsilateral EDL fibres. It thus appeared possible that nerve injections of colchicine were acting systemically, and indeed this was shown to be the case when similar denervation-like changes were caused bilaterally after intraperitoneal (I.P.) injections of the same dose of colchicine.</p> <p>Although thesis results showed that colchicine was acting systemically, it was still possible that it, was effective by blocking neuronal transport in the muscle nerves. Axoplasmic transport was therefore measured in the sciatic nerve of rats, following colchicine injections both systemically and into the contralateral nerve. At no time was there any evidence of an impairment of transport, even though both EDL muscles contained fibres which displayed extrajunctional sensitivity to ACh. lt was concluded that extrajunctional sensitivity to ACh following colchicine injections was not due to an interference with axoplasmic transport.</p> <p>In the injected sciatic nerve, an impairment of transport was consistently observed and this generally was associated with a detectable paresis and a small percentage (5-7%) of apparently denervated EDL muscle fibres. In addition, the indirectly evoked tetanic tensions produced by the ipsilateral EDL muscles decreased in comparison to the contralateral EDL muscles, commencing about 2 days following the sub-epineural injection of colchicine. From these observations, it was suggested that the demonstrable impairment of axoplasmic transport in the ipsilateral sciatic nerve caused the impairment of neuromuscular transmission on that side.</p> <p>Colchicine was also chronically applied to rat sciatic nerves by the use of drug-impregnated silicone rubber nerve-cuffs. This procedure prevented the systemic effect of colchicine, but the mechanical presence of the cuff resulted in a usually transitory and variable block of impulse conduction in the nerve. Since muscle inactivity per se also leads to changes previously believed to be due to denervation, the changes that were observed with colchicine-cuffs did not constitute unequivocal evidence supporting neurotrophic effects on the ipsilateral EDL muscle fibres.</p> <p>It was concluded from the results presented in this thesis that the results of experiments involving colchicine injections or drug-impregnated nerve-cuffs do not unequivocably support the existence of a neurotrophic control of skeletal muscle fibre properties.</p> <p>Finally, preliminary experiments in which colchicine was used in association with denervation indicated that the most likely mechanism of action of colchicine was a direct one on the muscle fibre membrane itself.</p> / Doctor of Philosophy (PhD)

<p>This research investigated absorption, distribution and utilization of Zn and Cu in developing piglets as a model for human premature infants, specifically focusing on the impact of diet (amount and ratios of minerals), drugs (dexamethasone) and stage of development on mucosal cell membrane Zn and Cu transport and body Zn and Cu storage. In 32 and 52 h old piglets, velocity of Zn transport across intestinal brush border membranes (BBM) was higher than in 10 and 20 d old piglets but occurred via a nonsaturable mechanism. Cu transport across BBM was greater in the 20 d group than in 10 d, 32 h and 52 h groups and occurred mainly via a saturable mechanism in 32 h, 10 d and 20 d groups. Plasma Zn and Cu were significantly lower but tissue Zn, Cu and metallothionein (MT) contents were higher in the younger groups than the 10 and 20 d groups. Competition for transport between elements may also be important in infants since Cu transport across BBM was suppressed by Zn but enhanced by Fe at the ratios of Zn:Cu and Fe:Cu which are similar to those in premature infant formulas. Further studies indicated that Fe mainly increases Cu binding to BBM. High dietary Zn intake did not alter Zn transport across BBM but induced Zn accumulation in the intestinal mucosa and liver in piglets. These results are valuable in determining appropriate upper limits for Zn, Cu and Fe contents in infant formula. Exogenous dexamethasone (DEX) when used therapeutically to induce lung maturation in premature infants may compromise Zn and Cu status. Zn influx across BBM was significantly greater but Zn efflux was significantly lower in DEX treated compared to control piglets. The effect of DEX on Zn influx was abolished by dietary Zn supplement. Intestinal Cu uptake was also enhanced by DEX treatment. However, DEX-induced mucosal uptake of Zn and Cu did not appear to result in increased net Zn and Cu absorption since intestinal MT was induced by DEX. This study provides evidence that DEX treatment alters Zn and Cu metabolism in early life. Zn and Cu status should be carefully monitored in premature infants receiving long-term DEX therapy.</p> / Doctor of Philosophy (PhD)

<p>A review of the available evidence relating herpes simplex type 2 (HSV-2) to cervical cancer in humans showed reasonable circumstantial evidence to suspect the participation of HSV-2 in the pathogenesis of the tumors. The best evidence was provided by seroepidemiological data, however, variations were observed among the data obtained from different populations. A mathematical model based on causal considerations and developed as an extension of a model used in chemical carcinogenesis was found to account for a large part of these variations. This model was found to reasonably describe the relation between the incidence rate of cervical cancer and the fraction of women with previous experience with the virus in different human populations. The fitting to the data suggested the existence of two kinds of etiologic factors involved in the pathogenesis of cervical cancer, one related to HSV-2 and another not related to the virus. The model described for average population values was expanded to age-specific data. This allowed the estimation of the age distribution of primary infection with HSV-2 (ϕ(v)), the distribution of the age of developing cancer unrelated to HSV-2 (Yᵤ(t)), the distribution of the age of developing cancer associated with HSV-2 (Yᵥ(t)), and the distribution of the time elapsing from the primary infection with HSV-2 up to the development of cervical cancer associated to HSV-2 (ψ(u)). The different shapes of Yᵤ(t) and Yᵥ(t) supported the existence of the two kinds of etiologic factors in the pathogenesis of the tumors. In addition, ψ(u) was found to correspond to relatively long times suggesting an important role for viral latency or viral reinfections in HSV-2 carcinogenesis. In view of the fact that some cervical cancers may not be related to HSV-2 the direct detection of HSV-2 genetic information in the cancer cells was considered fundamental for the correct assessment of the viral participation in the pathogenesis of the tumor. Large inconsistencies were found among reports of studies designed to detect HSV-2 genes in cancer cells. As a consequence a new approach was explored. The direct retrieval of viral endogenous genes was attempted from cells biochemically transformed by the virus. Contrary to reports from other systems, rescue was not detected. The frequency of rescue products was estimated as a value below 7x10^-17. These results suggested that rescue experiments which may be suitable for the detection of latent HSV, are not sensitive enough for the exposure of subgenomic viral sequence present in transformed cells. As a requirement for the rescue experiments, a new technique was developed to quantitate and clone HSV expressing the viral thymidine kinase in stocks predominantly lacking virus able to express this enzymatic activity.</p> / Doctor of Philosophy (PhD)

<p>Pulmonary structure and function were compared in healthy adult horses and cows to examine the hypothesis that differences in pulmonary function between two species of similar size were related to differences in pulmonary structure. The functional residual capacity and tidal volume were higher in the horses than in the cows while the respiratory rate was higher in the cows. The pattern of air flow differed between the two species, and flow rates were higher in the cows. The dynamic compliance was higher in the horses while the work of breathing was higher in the cows. Separation of the lower pulmonary system (trachea to pleural cavity) from the total pulmonary system (nares to pleural cavity) indicated that most of the resistance and work of breathing were in the upper airways (nares to trachea), and there were no differences in the mechanical indices of pulmonary function in the lower pulmonary system between the two species. To more adequately characterize the lungs, horses and cows were anaesthetized and pressure-volume manoeuvres were performed to determine the lung capacities and quasistatic compliance. The lung capacities were significantly greater in the horses, but the lung compliance did not differ significantly between the two species. Quantitative and qualitative observations revealed few significant differences between the structure of the lung parenchyma in horses and cows. The obvious structural differences were in the size and shape of the lungs, the branching pattern of the airways, and the amount and distribution of the interlobular septa. During the measurements of pulmonary mechanics in the standing animals there were several anomalous observations and examination of the pressure recordings revealed phase lags between the total and lower pulmonary systems. It would appear that it is invalid to assume that inertia is a negligible factor in pulmonary function in large mammals such as the horse and the cow, due to the large abdomen in these species. The size and shape of the abdomen has probably been a significant factor in the evolution of lung structure in horses and cows, and abdominal movements are important in determining the breathing patterns in these two species.</p> / Doctor of Philosophy (PhD)

<p>Increased platelet aggregation and Telease of granule contents in response to ADP,. epinephrine, and collagen, increased platelet factor 3-availability induced by ADP or collagen, increased platelet turnover and shortened template bleeding times in patients with type II- and type IV-hyperlipidemia have been reported from several laboratories. Furthermore, studies in experimental animals and in man have indicated that diet-induced elevation of blood Iipid levels may be associated with increased platelet adhesiveness, increased platelet factor 3-availability in response to ADP, and increased platelet turnover. Recently, evidence has been reported that in vitro exposure of pIatelets to increased levels of cholesterol in the medium may be associated with an increase in platelet cholesterol/phospholipid ratio and platelet aggregability. It has also been demonstrated that changes in phospholipid-fatty acid composition of the platelets may occur as a result of diet-induced changes in phospholipid-fatty acid composition of the plasma. Although long chain saturated fatty acids are known to induce or enhance platelet aggregation in vitro, uptake of free fatty acids appears to occur only when the concentration of free fatty acids exceeds the plasma albumin free fatty acid binding capacity. Phospholipids constitute a major proportion of the platelet membranes. Phosphatidic acid and the phosphoinositides appear to be involved in platelet shape change, aggregation, and the platelet release reaction. Certain phospholipids in platelets, when exposed, are powerful accelerators of blood coagulation reactions in vitro.</p> <p>The primary aims of this study were to investigate a) the possibility of a direct exchange of phospholipids between plasma and platelets in vitro; and b) the possibility that phospholipid exchange could result in changes in platelet function.</p> <p>In preliminary studies evidence was obtained to indicate that sodium pentobarbital used as anesthetizing agent in the blood collection procedure in rabbits inhibited platelet adherence to collagen-coated surfaces and the platelet release reaction when the compound was added to platelets in vitro. This effect of SPB appeared not to be related to SPB-induced suppression of platelet energy production but rather due to platelet membrane stabilization. The inhibitory effect of SPB on platelet function was readily reversible and SPB-induced anesthesia in rabbits was not found to cause measurable inhibition of pIatelet function.</p> <p>A sensitive and practical assay for collagen-induced platelet factor 3-availability; using a modified Stypven-clotting time method, was elaborated. During these methodological studies, new evidence with respect to the mechanism of platelet factor 3-availability was obtained. The findings indicated that with platelets from rabbits, pigs and humans platelet factor 3 did not become available in association with platelet shape change or platelet aggregation, but was rather closely linked to the platelet release reaction. However, a small amount of lysis was consistently observed in association with the platelet release reaction and it could not be clearly differentiated whether increased platelet factor 3 availability was truly a result of fusion of granule membranes with the external plate let membrane during the release reaction or a result of release-associated platelet damage.</p> <p>When hyperlipidemia was induced in rabbits by feeding them a diet enriched with egg yolk for a minimum period of 16 weeks, platelets isolated from the blood of these rabbits and resuspended in Tyrode-albumin solution showed a significant increase in collagen- or thrombin-induced aggregation and serotonin release, but not in aggregation induced by ADP. Collagen-induced platelet factor 3-availability was also greater with platelets from hyperlipidemic rabbits compared with platelets from control rabbits. There were no significant differences between the diet and control groups in platelet aggregation induced by ADP, collagen, or thrombin or serotonin release induced by collagen or thrombin in citrated platelet rich plasma. With washed platelets, there was no correlation between the percentage increase in aggregation induced by collagen or thrombin and the plasma cholesterol or triglyceride levels in hyperIipidemic or control rabbits. When washed normal rabbit platelets were resuspended in citrated plasma from a hyperlipidemic rabbit, their response to aggregating or release-inducing stimuli was not significantly different from that observed when platelets from normal rabbits were resuspended in normal plasma. When washed normal were incubated for 30 minutes in hyperlipidemic plasma at 37°C, isolated from the plasma and resuspended in Tyrode-albumin solution, their response to aggregating or release-inducing stimuli was not significantly different from that observed with normal platelets incubated in normal plasma. These findings, thus, support previous reports that diet-induced hyperlipidemia in rabbits may induce an intrinsic functional alteration of the platelets which cannot be readily reproduced by short-term in vitro exposure of platelets to hyperlipidemic plasma.</p> <p>The possibility of a direct exchange of preformed phospholipids between pIatelets and plasma was explored by adaptation of a model previously used in the study of phospholipid exchange in erythrocytes. Washed rabbit platelets were resuspended in plasma in which all of the major phospholipids had been isotopically labeled by injection of ³²PO₄ into rabbits. At certain intervals during a 6-hour incubation at 37°C, aliquots were removed from the incubation mixture and the platelets were isolated and subjected to lipid extraction and phosphilipid analysis. A continuous rise in platelet non-lipid-bound and lipid-bound radioactivity was observed-throughout the incubation period. Two platelet phospholipids, lecithin and lysolecithin, were significantly labeled, whereas little or no labeling of the other phospholipids was found. There was no detectable change in total or individual platelet phospholipid content. At 6 hours, 4% of total platelet phospholipids, 43% of platelet lysolecithin, and 7% of platelet lecithin were labeled. Platelets incubated in plasma from rabbits with diet-induced hyperlipidemia took up and incorporated significantly more label into their phospholipids than did platelets in normal plasma but the pattern of labeling (PC + LPC) was similar to that observed normal plasma. Labeling of both platelet lysolecithin and lecithin could be explained by uptake and metabolism of plasma lysolecithin by platelets. However, labeling of platelet lecithin could at least in part be the result of direct exchange of this phospholipid with the plasma. This is the first demonstration that platelets take up and metabolize endogenous plasma lysolecithin and, possibly, directly exchange lecithin and that these processes may be significantly accelerrated in hyperlipidemic plasma. Uptake and incorporation of endogenous plasma lysolecithin by platelets and, possibly, direct exchange of platelet lecithin appear to be important for platelet membrane phospholipid renewal and could be an important mechanism by which plasma lipids could modify platelet membrane phospholipid-fatty acid composition and, possibly, platelet function.</p> <p>In view of the possibility that lysolecithin might be an important link in the interaction between platelets and plasma lipo-proteins and apparent contradictions in the data reported from other laboratories, the effects of lysolecithin on aggregation, serotonin release, shape and lysis of rabbit, pig, or human platelets in platelet-rich plasma or Tyrode-albumin solution were examined during prolonged incubation. Lysolecithin added to citrated or heparinized PRP from humans or rabbits at a final concentration above 100 μM, caused instantaneous inhibition of platelet aggregation induced by ADP, epinephrine (human PRP only), collagen or thrombin. The inhibitory effect of lysolecithin was found to be partially reversible over a period of 60-90 minutes. Lysolecithin at final concentrations above 30 mM also caused inhibition of ADP-, collagen- and thrombin-induced aggregation and collagen and thrombin-induced release of serotonin in suspensions of rabbit, pig, or human platelets. With washed platelets, the inhibitory effect not only rapidly disappeared, but was followed by transient potentiation of aggregation and serotonin release. This potentiating effect of lysolecithin was most pronounced when thrombin was used as stimulus. Both inhibition and potentiation were observed at concentrations of lysolecithin that did not cause a significant change in platelet shape or loss from platelets of lactic dehydrogenase. Inhibition and potentiation were also observed when plafelets were added to suspending medium containing lysolecithin, although considerably higher concentrations of lysolecithin were required under these conditions. Potentiation was not observed when lysolecithin was added to citrated or heparinized rabbit or human PRP or to washed rabbit platelets suspended in a medium containing 4% bovine serum albumin. It seems Iikely that some or all of the observed effects of lysolecithin on platelet function are due to structural modification of the platelet membrane insufficient to result in gross membrane damage of platelet lysis. In addition, the results of experiments using ¹⁴C-Iysolecithin seemed to indicate that the reversal of inhibition and the potentiating effect of lysolecithin on platelet function may at least in part be related to its rapid uptake and metabolism by the platelets. Whether plasma lysolecithin plays a role in the regulation of platelet function under normal conditions and/or in hyperlipidemia and whether acute changes in plasma lysolecithin concentration (e.g. in response to heparin administration) can acutely modify platelet function remains speculative.</p> <p>Lysophosphatatidylethanolamine and Iysophosphatidylserine also caused inhibition of platelet aggregation and serotonin release in suspensions of washed rabbit and human platelets induced by ADP, collagen or thrombin. With washed pig platelets lysophosphatidylethanolamine inhibited platelet aggregation and the platelet release reaction whereas lysophosphatidylserine induced platelet aggregation and serotonin release. All of these effects on platelets were observed at concentrations of the lysophospholipids which did not cause gross platelet membrane damage or platelet lysis. The differences in the effects of these lysophospholipids on platelet function could be related to marked differences in their fatty acid composition.</p> <p>The results of this study contribute to our knowledge of the mechanisms of platelet factor 3-availability, platelet membrane phospholipid renewal and the role of plasma lipids in the modification of platelet phospholipid and phospholipid-fatty acid composition and platelet function.</p> / Doctor of Philosophy (PhD)

<p>Macrophages obtained from animals infected with intracellular parasites are activated with respect to their ability to inhibit the replication of the parasite. While normal rabbit macrophages support the replication of vaccinia virus at levels of 2 to 3 logs, macrophages obtained from the peritoneal cavity of rabbits infected 9 to 12 days earlier with vaccinia virus are activated and will not support virus replication. The fate of vaccinia virus in activated rabbit macrophages was studied in order to characterize the abortive infection of the virus within the activated macrophage. Vaccinia virus adsorption was measured using radioactively labelled virus and was similar with both normal and activated macrophages but was lower than, the amount of virus which adsorbed to Vero cells. Maximum adsorption took place during the first 10 minutes of incubation. A significant amount of the vaccinia virus which had adsorbed to the cells eluted from the cells during further incubation. However, the virus elution curves for activated and normal macrophages were similar. Virus uncoating was measured by infecting with ³H thymidine labelled vaccinia virus and then detecting DNase sensitive, TCA soluble counts. Vaccinia virus was able to uncoat to a similar degree in both activated and normal macrophages. Maximum virus uncoating took place one to four hours after adsorption and was approximately 55% of the virus which was adsorbed to the cells. DNA synthesis in vaccinia virus infected cells was detected by pulse labelling with ³H thymidine. A burst of DNA synthesis at 3 to 6 hours after infection took place in both activated and normal macrophages infected with vaccinia virus as well as infected Vero cells. The pattern of vaccinia virus antigen production in activated and normal macrophages was identical as detected by immunofluorescence and immunodiffusion. Autoradiographs of SDS-PAGE gels of lysates of infected cells pulsed with ³H amino acids demonstrated that most of the polypeptides formed within the infected macrophages were identical. However, at least three polypeptides present in the activated macrophages infected with vaccinia virus were absent in the infected normal macrophages and at least one polypeptide present in the virus infected normal macrophages was absent in the virus infected activated macrophages. Pulse chase experiments failed to demonstrate that the differences in polypeptide synthesis in activated and normal macrophages infected with vaccinia virus were due to differences in posttranslational cleavage. Lack of virus particle production in activated rabbit macrophages infected with vaccinia virus was the major detectable defect in the viral replicative cycle. Virus particles were defected by centrifuging on a continuous sucrose gradient cell lysates of virus infected cells labelled with radioactive thymidine or amino acids. No virus particles with the size and density of vaccinia virions were detected in lysates of activated macrophages infected with vaccinia virus. Virus particles were present in normal macrophages and Vero cells after vaccinia virus infection.</p> <p>It appears that the inhibition of production of infectious virus in activated macrophages is mediated by mechanisms other than those induced by interferon. Vaccinia virus DNA and protein were synthesized in activated macrophages. This is in contrast to numerous previous studies which have shown that no vaccinia viral DNA or protein is synthesized in interferon treated cells. Retreatment of normal rabbit macrophages with tissue culture interferon (type I) did not block the replication of vaccinia virus. Pretreatment of normal macrophages with serum from a poly (I)-poly (C) injected rabbit reduced the replication of vaccinia virus but the characteristics of the abortive infection appeared different than in activated macrophages. Therefore, it appears that interferon is not involved in the inhibition of viral replication by activated macrophages.</p> / Doctor of Philosophy (PhD)

<p>Congenital hip disease (CHD), a condition in which the head of femur may be partially or completely separated from the acetabulum, is detected in one to 4 infants per 1,000 live births. At birth the hip joint is cartilaginous and underdeveloped. In cases of CHD the acetabulum is shallow, the femoral head is small and aspherical, ligament of the head of femur (LHF) is elongated, and the femoral angles are abnormal. These are measureable characteristics. However, no previous studies have given quantitative data for several dimensions of this joint in a sample of fetuses, at regular intervals of age, throughout the fetal period.</p> <p>The pattern of normal growth and development was studied in 280 hip joints from 140 fetuses between 12 weeks and term. Measurements were taken of acetabular depth, diameter, femoral head diameter, the length and width of the LHF, and the femoral torsion and neckshaft angles. Histological studies of ossification and labrum structure were undertaken on a number of acetabula at intervals throughout the fetal period.</p> <p>Acetabular depth was shown to be the slowest growing variable at the hip joint. The study confirmed previous reports that the acetabulum tends to become shallower towards term but suggested that the greatest amount of change in shape may occur early in the fetal period. In a sample of femoral heads a tendency for the head to become less spherical with age was revealed. With the exception of the neckshaft angle all variables demonstrated a moderate to high positive correlation with age. Variables were best fitted by a regression model which included a polynomial on age. Only for left LHF length and right LHF width was growth a simple interest function. The velocity of growth for ail dimensions, except the neck-shaft angle, was highest between 12 and 18 weeks. After this period a deceleration in the growth rate was noted, most noticeable in acetabular depth.</p> <p>No significant differences were detected between males and females, or between the right and left sides. This indicates that the higher incidence of CHD in females, and the greater involvement of the left hip must be explained by factors other than the growth of cartilage and bone.</p> <p>The LHF was shown to be variable in shape throughout the fetal period. It was not a distinctly linear structure in normal or abnormal hip joints. A strong correlation with acetabular dimensions was not evident.</p> <p>Booth femoral angles demonstrated variability throughout the fetal period and values reported are lower than those currently accepted for the newborn. Since the angles demonstrated poor to moderate correlation with the other hip dimensions, neither angle alone appears to provide a useful indicator of normal development. A change in the orientation of the lesser trochanter with age, correlated with the increase in torsion was observed. This is considered pertinent to the reading of femoral angles on radiographs.</p> <p>In 65 hip joints from 46 fetuses variation from descriptions of normal hip joint morphology was detected. Measurements of these joints did not differ significantly from those of normal hip joints. It is suggested that a number of these variants are microforms of CHD. An unexpected number of variants and abnormal socket features were localized to the anterosuperior quadrant and anterior socket wall.</p> <p>Comparison of data from abnormal joints (18) with data from the normal growth study permitted a more precise evaluation of the abnormality. From this comparison underdevelopment of apparently "normal" joints was detected.</p> / Doctor of Philosophy (PhD)

<p>To test the error catastrophe theory of cell senescence, we have developed a cell-free protein synthetic system from human diploid fibroblasts capable of translating both the synthetic mRNA, poly (u), and endogenous cellular mRNA in vitro. We have measured the fidelity of poly(U)-directed protein synthesis in extracts from a variety of cells. The results contradict the error catastrophe in several ways: - cells from subjects suffering maladies of premature aging, progeria and Werner Syndrome, exhibited error frequencies within the normal range. - cells from an old donor did not have elevated error frequency of protein synthesis. - early-passage (young) normal cells had error frequencies higher than those of late-passage (old) cells. The error frequency in one normal strain of fibroblasts declined throughout its tissue culture lifespan. Cells maintained in the post-mitotic (terminal) state, in which there was no cell selection, exhibited a constant error frequency over 16 weeks. In a series of experiments using clonal populations of cells, it became evident that the decline in error frequency as a function of passage was dependent on the clonal heterogeneity of the mass culture. In no experiment, however, could any correlation between the error frequency of protein synthesis and cellular growth parameters be made. Overall, the senescence characteristic of human diploid fibroblasts was independent of the measured fidelity of protein synthesis in vitro.</p> / Doctor of Philosophy (PhD)

<p>The changes in the physiological properties of the sinoatrial node which follow a programme of exercise training have been examined in the rat. The influence on the intrinsic sinoatrial frequency (ISF) of altered levels of autonomic nervous activity accompanying the physical training has been studied in two series of experiments combining daily pharmacological agents with ten weeks of treadmill exercise. The first series (Study I) examined the influence on the ISF of the increased parasympathetic activity reported in the trained state by blocking with atropine or stimulating with carbachol the receptors of the parasympathetic system in the heart. The second series (Study 2) examined the influence on the ISF of the increased sympathetic activity which occurs during exercise by stimulating with noradrenaline or isoprenaline, or blocking with propranolol the receptors of the sympathetic system in the heart. The content of potassium (K) and sodium (Na) in the plasma and myocardium were measured in all animals at the completion of the ten week period, and an estimate of electrophysiological changes was derived from the plasma to myocardial K⁺ ratio (K⁺o/K⁺i). In addition, the chronotropic response of the sinoatrial node to acetylcholine and noradrenaline was examined in vitro for all animals in Studies 1 and 2. The chronotropic responses of the sinoatrial node to exercise and noradrenaline were further explored in the in vivo experiments of Study 3.</p> <p>In Study 1, ISF was reduced with training. Blockade of the parasympathetic receptors with atropine also reduced ISF, and the effects of exercise and atropine appeared to be additive. Stimulation of the parasympathetic receptors with carbachol had little effect on ISF. In Study 2, stimulation of the sympathetic receptors with either noradrenaline or isoprenaline caused a reduction in ISF, but when combined with exercise, no reduction occurred. Sympathetic receptor blockade with propranolol reduced ISF, and propranolol plus exercise also reduced ISF. It appears that the exercise-induced reduction of ISF was not a consequence of increased parasympathetic activity in the trained state, or of increased sympathetic activity during exercise. The K⁺o/K⁺i was reduced with training. This was consistent with a more negative maximum diastolic repolarization, and a reduced ISF. The K⁺o/K⁺i of the drug treated groups indicated that ISF was probably reduced through factors other than a more negative maximum diastolic repolarization. No difference was found in the sensitivity of the atria of any group to acetylcholine in vitro. Increased chronotropic sensitivity to noradrenaline was found in vitro in the two chronically noradrenaline treated groups. The maximum response to noradrenaline was reduced in vitro. The maximum cardiac frequency in response to either exercise or noradrenaline infusion in vivo was also reduced. This reduction of maximum frequency in both the atria and the intact heart was observed to be positively correlated with the ISF, thus those hearts with a low ISF obtained a low maximum frequency. This observation led to the proposal that electrophysiological properties of the sinoatrial node which establish the ISF are also involved in imposing a limit on the maximum sinoatrial frequency.</p> / Doctor of Philosophy (PhD)