Fetal tissue repair and wound healing

The fibrosis and scar formation that characterize adult wound healing are also the cause of clinical problems; scar contracture, hypertrophic scar, and pulmonary and hepatic fibrosis are only a few examples. Studies of fetal wound healing can provide an insight into the initiation and regulation of a scarless repair process akin to regeneration. Studies of fetal repair have already suggested mechanisms that might favorably alter adult healing. Topical application of hyaluronic acid to wounds in adult diabetic rats leads to enhanced epithelial migration. It has been recognized that the addition of TGF-beta to fetal wounds causes an adultlike healing response with fibrosis and inflammation. A subsequent study using neutralizing antibody to TGF-beta in adult wounds showed enhanced healing with a more normal dermal architecture with fewer macrophages, fewer blood vessels, and less collagen. As our understanding of regenerative tissue repair increases, the opportunities to modulate adult fibrotic conditions should expand.     

The in vivo effect of hyaluronan associated protein-collagen complex on wound repair

Fetal skin wounds heal without scarring, however the underlying mechanisms remain unknown. Immunohistochemical staining and biochemical studies indicate the deposition of a collagen repair matrix that is highly organized. We have previously described a unique hyaluronan associated protein-collagen complex (HA-PC) profile present during the period of scarless healing in the sheep fetus. In this study, we examined the biologic activity of this HA-PC in an in vivo model of adult rat wound repair. A total of 84 incisional and 84 excisional wounds were examined by histology, TGF-beta immunocytochemistry and computer planimetry (excisional wounds only). Planimetry of the excisional wounds demonstrated the mean wound area remaining at day 1 was 88.7% for the control and 63.6% for the treated (p<0.01). At day 2, mean wound area was 81.5% for the control and 63.6% for the treated (p<0.01). At day 4, mean wound area was 56.6% for the control and 41.9% for the treated (p<0.01). At day 7, mean wound area was 26.9% for the control and 16.8% for the treated (p<0.01). At day 14, mean wound area was 7.9% for the control and 3.4% for the treated (p<0.05). Collagen organization was judged to be greater in the treated compared to control wounds, with a mean organization score of 2.3 vs. 1.9 (p=0.0596; Wilcoxon Signed Rank Sum Test). There were significantly more neutrophils at the wound margin of the treated compared to control wounds, 4.0 vs. 2.7 (p=0.038; Paired Two Tailed Student's t-Test). There was no difference in the number of microphages at the wound margin of the treated compared to control wounds, 6.15 vs. 6.0 (p>0.05). TGFbeta1 and beta2 staining was decreased whereas TGFbeta3 staining was increased in the HA-PC treated wounds. These results suggest that compared to control wounds HA-PC treated wounds heal more quickly, with more organized collagen, more neutrophils at the wound margin and increased TGFbeta3 expression. Furthermore, these data suggest that the manipulation of scarring in adult wounds is possible by the addition of proteins present in fetal skin.     

Dimensions of schizophrenia in the elderly. Correlations with cognitive and motor symptoms

Wound contraction is thought to be independent of site, and circular full-thickness skin wounds are though not to contract completely. To verify these statements four circular full-thickness skin wounds were created on each side of eight pigs and randomised to treatment with either split-thickness skin grafts, or healing by secondary intention under a hydrocolloid dressing. Time to healing, contraction, and final scar shape were evaluated. The median healing time was 12 days (range 6-18) in the grafted wounds and 30 days (range 15-45) in the secondarily healing wounds. There were significant differences in healing time between the different sites on the pigs. In the secondarily healing group, medial-caudal wounds healed in 21 (15-21) days compared with lateral wounds which healed in 36 (21-45) days (p < 0.005), while no differences were found in the grafted group. There was a clear relationship between site and contractility and shape of the scars in both treatment groups. Scars located on the lateral-caudal aspect of the pig were predominantly round and contracted only slightly. Scars located on the lateral aspect of the pig tended to be oval. Contraction was greatest in the medial scars and least in the lateral scars. Median contraction was 33% (range -2-63) in skin grafted wounds and 64% (range 42-82) in secondarily healed wounds. This randomised experiment showed that extent of wound contraction is dependent of site, and that circular wounds do heal with contraction.     

Effect of sciatic nerve transection or TTX application on enzyme activity in rat spinal cord

BACKGROUND/PURPOSE: Treatment of several congenital anomalies is frequently hindered by lack of enough tissue for surgical reconstruction in the neonatal period. The purposes of this study were (1) introduction of a novel concept in perinatal surgery, involving minimally invasive harvest of fetal tissue, which is then processed through tissue engineering techniques in vitro while pregnancy is allowed to continue, so that, at delivery, the newborn can benefit from having autologous, expanded tissue promptly available for surgical implantation at birth; (2) analysis of the progress of an engineered fetal skin graft with time, after implantation in the neonate; and (3) study of the effects of current tissue engineering techniques on fetal keratinocytes and fetal dermal fibroblasts. METHODS: Ten 90- to 95-day-gestation fetal lambs underwent surgical creation of two large paramedian excisional skin defects on the posterior body wall. Subsequently, fetal skin specimens no larger than 1.5 x 1.5 cm were videofetoscopically harvested. Fetal keratinocytes and dermal fibroblasts were then separately cultivated and expanded in vitro for 45 to 50 days, resulting in a total of approximately 250 to 300 million cells. Seven to 10 days before fetal delivery, all cells were seeded in two layers on a 16 to 20-cm2, 3-mm thick biodegradable polyglycolic acid polymer matrix. One to 4 days after delivery, the autologous engineered skin was implanted over one of two previously created skin defects. The second skin defect region received an absorbable polymer scaffold without cells as a control. If necessary, the original skin wounds were further amplified before implantation. Each animal provided at least one time-point for histological analysis of both types of repair through excisional biopsies performed at weekly intervals, up to 8 weeks postimplantation. Normal skin specimens were also used as controls. RESULTS: Fetal and neonatal survival rates were 100%. Based on previous postnatal skin engineering studies, fetal dermal fibroblasts multiplied significantly faster in vitro (approximately fivefold) than expected. Fetal keratinocytes multiplied at expected postnatal rates. The engineered grafts induced faster epithelization of the wound (partial at 1 week and complete between 2 and 3 weeks postoperatively) than did the acellular ones (partial at 3 weeks and complete between 3 and 4 weeks postoperatively). Analysis of skin architecture showed a higher level of epidermal organization and less dermal scarring in the wounds that received the engineered, cell-implanted polymer scaffold. CONCLUSIONS: (1) Videofetoscopically assisted fetal tissue engineering is a viable method for obtaining expanded autologous tissue for prompt surgical reconstruction at birth. (2) Fetal skin can be expanded and engineered in vitro at faster rates than expected postnatally, with current tissue engineering techniques. (3) Engineered autologous fetal skin induces a faster and more organized healing of neonatal skin defects than that observed with second intention. This concept may prove useful for the treatment of certain human neonatal conditions such as giant neoplasias, ectopia cordis, and other body wall defects.     

Tissue repair in the fetal intestinal tract occurs with adhesions, fibrosis, and neovascularization

Cutaneous wound healing in the fetus can occur in a nonfibrotic, regenerative manner. However, other fetal tissues such as bone and stomach heal with scar formation. In light of potential ramifications for adult hollow visceral scarring (biliary and intestinal strictures), this study was undertaken to determine if tubular visceral tissue repair in the fetus is regenerative or fibrotic. Fetal rabbits underwent laparotomy on day 24 of gestation, during which a controlled intestinal enterotomy was created and suture repaired immediately using microsurgical techniques. Maternal rabbits and adult male rabbits also underwent enterotomy and repair. After 5 days all animals were sacrificed and the wounds analyzed histologically by a pathologist in a blinded fashion. All animals demonstrated a similar degree of peri-intestinal adhesion formation. Fetal and maternal wounds contained fibroblastic and smooth muscle cell proliferation, mild inflammatory infiltration, and new blood vessel formation. The male adult wounds demonstrated a more pronounced fibrovascular healing response. Immunohistochemical staining for CD31 (endothelial cell marker) was quantitated on a scale of 0 to 4+, indicating degree of neovascularization. The mean scores for the fetal and maternal groups were similar (1.70 +/- 0.68 and 1.23 +/- 1.07 respectively), but were significantly greater for male adults (2.93 +/- 0.12; p = 0.001 by analysis of variance). The results of this study indicate that hollow visceral tissue repair in the fetal rabbit intestine occurs in a similar fibrotic manner as adult healing. This provides further evidence that regenerative healing in the fetus is not ubiquitous. Differences in the degrees of fibrosis and neovascularization between adult male and pregnant female wounds deserve further investigation.     

Influence of biosynthetic human growth hormone on the biomechanical strength development in skin incisional wounds of diabetic rats

The influence of biosynthetic human growth hormone (b-hGH) on the mechanical strength development of skin incisional wounds in diabetic rats has been investigated after 4, 7 and 21 days of healing. Diabetes caused decreased mechanical strength of the wounds. After 21 days of healing, the diabetic animals treated with b-hGH from 7 days before wound infliction had an increased maximum load (14%) compared with the untreated diabetic animals. After 4 and 7 days of healing, no differences in the biomechanical strength of diabetic wounds with and without b-hGH treatment were found. The diabetic animals receiving b-hGH showed a significant increase in body weight compared with the untreated diabetic animals. In the diabetic animals, the blood glucose concentration was increased by treatment with b-hGH. In conclusion, treatment with b-hGH from 7 days before operation can counteract the reduced mechanical strength of skin wounds in diabetic rats on day 21.     

Hyaluronic acid inhibits fetal platelet function: implications in scarless healing

Platelets are important for the initiation of inflammation in adults, but the role of fetal platelets in fetal wound healing is unclear because fetal dermal wounds heal with a minimal inflammatory response and lack of excessive scarring. Because fetal tissue is abundant in glycosaminoglycans (GAGs), predominantly hyaluronic acid (HA), this study was designed to test the hypothesis that HA inhibits the reactivity of platelets and thus contributes to the minimal scarring characteristic of fetal tissue repair. Platelets were isolated from 10 fetal pigs at day 80 of gestation (term, 115 days) and exposed to 0.5 mg/mL of arachidonic acid, an agent shown in prior studies to evoke maximal aggregation and degranulation of fetal platelets. The ability of HA at 0.1 and 0.5 mg/mL to inhibit this response was determined. The presence of HA resulted in a dose-dependent reduction in platelet aggregation at 180 seconds (control, 99.7 +/- 0.3%; HA [0.1 mg/mL] 91.7 +/- 3.8%; and HA [0.5 mg/mL] 48.5 +/- 9.0%; P < .005 v control). The onset of aggregation was also significantly delayed by 0.5 mg/mL of HA (13.5 +/- 2.5 seconds) compared to control (2.9 +/- 0.7 seconds), P < .05. No significant diminution of platelet aggregation could be achieved by the addition of other GAGs at similar concentrations. HA also significantly impaired the release of platelet-derived growth factor (PDGF)-AB from fetal platelets. The authors conclude that HA, the predominant GAG in fetal dermal matrix, inhibits platelet aggregation and cytokine release. This inhibition of platelet aggregation and resultant inflammatory response may explain, in part, the minimal inflammation and scarless healing characteristic of fetal dermal repair.     

Adrenal hemangioma: review of the literature

Initially thought to act as tissue replacement, cultured epithelial allografts are now known to work by providing a potent stimulus for healing. In a similar fashion, we believe that traditional autografts may also provide a stimulus to help heal chronic wounds, thus acting as pharmacological agents for healing. We attempted to assess the possibility of augmenting the stimulatory properties of donor skin by initiating the healing process in the donor region prior to grafting. This was accomplished by pre-wounding the donor area 3 days prior to harvesting the donor skin. We compared these 'pre-wounded' grafts to those harvested immediately. Two patients underwent punch grafting for chronic leg ulceration. Half of the ulcer was grafted with donor skin harvested from an area that was pre-wounded and the other half from freshly harvested skin. We evaluated each for improvement of granulation tissue and degree of edge effect (migration of the previously dormant wound edges). All the grafts did well. There was marked improvement in granulation tissue in the ulcer bed after grafting, and the obvious presence of an edge effect. The edge effect was increased on the site where the pre-wounded grafts were placed. It may be possible to augment the growth stimulatory properties of donor skin. This may offer therapeutic options in patients with chronic wounds.     

Psychologic factors involved in the decision to undergo reconstructive surgery after burn injury

We have developed a new in vivo model for the study of fetal wound healing. Fetal ICR mice (total gestation, 21 days) received a full-thickness incisional wound in the hind limb at gestational day 14 (N = 100). The wound was made with a 28.5-gauge needle that was passed transplacentally into the amniotic cavity. The wounds were analyzed histologically on postoperative days 0, 1, 3, and 5 by hematoxylin-eosin and Mallory's trichrome stains. Once the wounding technique was mastered, the overall mortality rate for this model was 20% by postwounding day 5. Each fetus healed their wound without scar by postwounding day 3. In 3 animals, 5 microliters of human transforming growth factor beta 1 (25 micrograms per microliter) was injected into the wound site, resulting in scar and an inflammatory cell infiltrate, indicating that the 14-day-gestation fetal mouse can be manipulated if necessary. This model offers the advantages of an in vivo system that can be studied at an early gestational age. Furthermore, it is inexpensive, easy to manipulate, and can be studied with commercially available murine probes.     

The regulation of collagen in fetal skin wounds: mRNA localization and analysis

The deposition of collagen in fetal skin wounds has been shown in several animal models. The authors used a radiolabeled RNA antisense probe, complementary to the mRNA for the alpha-1 chain of human procollagen type I, to assess regulation of this collagen species in fetal and adult rabbit wounds. Dorsal skin wounds were placed on fetal and maternal animals at the beginning of the third trimester, and were harvested 3, 5, and 7 days later. In situ RNA/RNA hybridization was performed on suitable specimens, and morphometric analysis was carried out with a computerized LECO image analyzer. Fetal wounds exhibited an inflow of mesenchymal cells that produced collagen type I at levels higher than the surrounding tissue; this activity was highest on days 3 and 5 after wounding. Adult wounds had increased fibroblast presence by day 7, producing collagen type I at levels higher than those of adjacent unwounded tissue. Morphometric analysis of the signal produced by in situ hybridization and of the number of cells producing the signal in a given field showed that fetal wounds appear to produce collagen type I by an increase in the number of cells in the area of the wound--not by induction of the gene for procollagen type I. In contrast, adult wounds had both fibroblast migration and induction of procollagen type I mRNA synthesis. These findings imply multilevel regulation of collagen production in the adult and posttranslational regulation in the fetus.     

Characterization of fetal ovine renal dysplasia after mid-gestation ureteral obstruction

OBJECTIVE: The etiology and pathogenesis of renal dysplasia are poorly understood. To characterize the histologic changes in fetal renal dysplasia, we studied a fetal ovine model of urinary obstruction. DESIGN: Animal study. ANIMALS: Seven fetal lambs, and other lambs of the same gestational age as controls. INTERVENTIONS: Unilateral ureteral ligation on fetal lambs at approximately 70 days' gestation (term for sheep is 145 days), during nephrogenesis. Kidneys were subsequently collected, examined histologically and characterized by immunohistochemical tests involving cytokeratin antiserum and a monoclonal antibody to alpha-actin. OUTCOME MEASURES: Histologic changes in ligated fetal lamb kidneys, based on comparison with normal fetal lamb kidneys. RESULTS: At near term (140 days' gestation), the ligated kidney showed distorted and less abundant renal parenchyma than a normal control kidney. Upon microscopic examination, the ligated kidney displayed marked architectural distortion of the outer cortex, with abundant interstitial fibrosis, primitive ductules and glomeruli, and cysts of varying sizes lined by squamous and cuboidal epithelia and surrounded by a loose mesenchyme. The renal medulla contained differentiated collecting ducts, which were structurally distorted and less abundant than in normal control kidneys. The proximal and distal tubule elements were primitive and markedly underdeveloped. Cytokeratin immunoreactivity was present in the collecting duct epithelium and in the cuboidal epithelium lining many of the cortical cysts. Smooth muscle alpha-actin immunoreactivity was localized in the cortical region of the kidney, which highlighted the abundance and disorganization of the undifferentiated mesenchyme and identified the fibromuscular collars of the primitive ductules of the cortex and the distorted collecting ducts of the medulla. CONCLUSIONS: These results highlight the histologic changes resulting from unilateral ureteral ligation in fetal lambs. This model is useful in the study of the pathogenesis of fetal obstructive renal dysplasia.     

Centralized echocardiogram quality control in a multicenter study of regression of left ventricular hypertrophy in hypertension

Deep skin wounds in the adult mammal close spontaneously by epithelialization, wound contraction, and scar synthesis. In previous wound healing studies, it has been unsuccessfully attempted to separate from each other the natural processes that close wounds. In this study, we attempted to isolate skin regeneration from spontaneous processes of wound closure using "island" grafts. A porous analog of the extracellular matrix, composed of a graft copolymer of type I collagen and chondroitin 6-sulfate, was seeded with uncultured autologous keratinocytes and served to induce regeneration of the dermis and the epidermis. Grafts of the copolymer, measuring 1 x 2 cm, were placed in the center of 5 x 6-cm wounds in guinea pigs. By day 14, the edges of the island grafts were clearly separated from the host epidermis and dermis by a distinct bed of granulation tissue. Histologic study of island grafts on day 14 showed that the copolymer grafts had largely degraded and that a new epidermis and dermis had been synthesized in its place. The thickness of the new epidermis increased as the density of cells seeded into the graft increased. No synthesis of epidermis or dermis was observed in the granulation tissue outside the perimeter of the island grafts. We conclude that island grafting allows the study of early events in skin regeneration in isolation from epithelialization, contraction, and scar synthesis.     

Estimation of the developmental age of the ovine fetus and lamb

Fifty-six fetuses and 33 lambs were obtained from a flock of ewes at set gestational intervals between 50 to 180 days after conception. The fetuses and lambs were killed, disected and the sizes and weights of a wide range of skeletal and soft tissues were measured. Five morphological parameters emerged as most suitable for the determination of normal foetal developmental age. By plotting the mean value and ninety-five per cent tolerance limits, the rates of growth and the variability of each parameter were studied. Crown-anus length is useful for determining fetal developmental age from 50 to 100 days gestation; brain weight, long bone length and the number of appendicular ossification centres can be used to determine fetal development age from 50 days gestation until term.     

Developmental regulation of gastric somatostatin secretion in the sheep

Gastric somatostatin (SRIF) regulates gastric acidity by inhibiting gastric acid and gastrin secretion. SRIF secretion is increased by gastric acidity and also directly by regulators of gastric acid secretion such as gastrin. This direct effect has not been described in the developing animal, nor have the roles of intermediaries such as histamine and gastric acidity been defined. The present study aimed to establish the regulatory role of gastrin and histamine during development on SRIF secretion and also to determine whether the effects of gastrin and histamine are independent of gastric pH. Pentagastrin and histamine were infused on separate occasions into fetal sheep, newborn lambs, and 28-day-old lambs. To determine the roles of endogenous histamine and gastric pH, ranitidine (a histamine-2 receptor antagonist) and omeprazole (a H+/K+ ATPase inhibitor) were coinfused with the agonists. Plasma SRIF and gastrin concentrations were measured by RIA. Pentagastrin stimulated SRIF secretion in the fetus after 131 days of gestation (term is 147 days), whereas stimulation by histamine was effective only after birth. The SRIF stimulatory effect of pentagastrin in 28-day-old lambs was abolished by ranitidine, which also reduced this effect in the adult sheep. This inhibitory effect of ranitidine was shown to be a result of blockade of stimulatory H2 receptors, because in the adult blockade of acid secretion with omeprazole failed to attenuate the response of histamine. These results indicate that in the fetus, gastrin receptors, but not histamine receptors, are functionally involved in the stimulation of SRIF secretion. After birth, both gastrin and histamine stimulate SRIF, but the effect of gastrin is mediated at least in part by the release of endogenous histamine. These responses occur independently of changes in gastric acidity, supporting the concept of a direct negative feedback between SRIF and gastrin.     

Late recurrence of a large post-infarction left ventricular pseudo-aneurysm. A case report

OBJECTIVE: To evaluate effects of treatment with a pulsed electromagnetic field (PEMF) on healing of open and sutured wounds, clinicopathologic variables, and CNS activity of dogs. ANIMALS: 12 adult female Beagles. PROCEDURE: Open and sutured wounds were created in the skin of the trunk of the dogs. Dogs were divided into 2 groups. One group received PEMF treatment and 1 group served as untreated (control) dogs. The PEMF-treated dogs received treatment twice a day starting the day before surgery and lasting through day 21 after surgery. Wounds were evaluated by use of tensiometry, planimetry, laser Doppler perfusion imaging, and histologic examination. Clinicopathologic variables and electroencephalographic tracings were also evaluated. RESULTS: Use of PEMF treatment resulted in significantly enhanced epithelialization of open wounds 10 and 15 days after surgery. Five days after surgery, wounds of control dogs had a negative value for wound contraction, whereas PEMF-treated wounds had a positive value. The PEMF treatment did not cause significant changes in short-term planimetric, perfusion, tensiometric, histologic, clinicopathologic, or electroencephalographic results. CONCLUSIONS: The PEMF treatment enhanced wound epithelialization in open cutaneous wounds and provided indications of early contraction without significant short-term changes in other variables.     

Keratinocyte growth factor-2 accelerates wound healing in incisional wounds

BACKGROUND: Keratinocyte growth factor-2 (KGF-2) also described as fibroblast growth factor-10 (FGF-10) is a newly identified member of the fibroblast growth factor family. KGF-2 is 96% identical to the recently identified rat FGF-10 and specifically stimulates growth of normal human epidermal keratinocytes. The present study was undertaken to examine the effects of topically applied KGF-2 in an incisional wound healing model. KGF-2 treatment resulted in an improvement in incisional wound healing as characterized by an increase in breaking strength, collagen content, and epidermal thickness. METHODS: KGF-2 was topically applied to linear incisions made in the dorsal skin of Sprague-Dawley rats. Biomechanical testing was done using an Instron tensiometer for breaking and tensile strength determinations. Wound collagen content was determined using the Sircol collagen assay. Epidermal thickness measurements were conducted using Masson's trichrome-stained sections of the wound. RESULTS: A single topical application of KGF-2 at the time of wounding resulted in an increase in wound breaking and tensile strength at Day 5 after wounding. Breaking strength of KGF-2-treated wounds was significantly higher compared with the buffer control (1 microgram, 222.1 +/- 13.5 g, P = 0.0007; 4 microgram, 248.7 +/- 15.4 g, P = 0.0001; 10 microgram, 247.2 +/- 21.9 g, P = 0.001; buffer, 141.0 +/- 9.7 g). Epidermal thickness and wound collagen content were significantly increased following treatment with KGF-2. CONCLUSIONS: Based on our findings, KGF-2 is a potent stimulator of wound healing as demonstrated by increased mechanical strength accompanied by an increase in wound collagen content. KGF-2 could be an important cellular mediator responsible for the initiation and acceleration of wound healing and may enhance the healing of surgical wounds.     

Acute phase toxoplasma abortions in sheep

Within 13 days of the experimental infection of 15 oestrus-synchronised ewes with 2000 sporulated oocysts of Toxoplasma gondii at 80 to 90 days of gestation 11 had aborted. The infection induced pyrexia and specific antibody in all the ewes. One ewe resorbed its fetus, 11 ewes aborted and three delivered, at full term, live congenitally infected lambs whose pre-colostral serum was antibody-positive. Tissues from the aborted fetuses and placentae from the live lambs were examined for toxoplasma infection by polymerase chain reaction (PCR) amplification of the B1 gene and by mouse inoculation. The live lambs were all shown to be infected by both methods, but there was no evidence of infection in any of the tissues from the acute phase abortions, suggesting that these fatalities occurred before the placenta or the fetus had been invaded by T gondii. Such toxoplasma-induced, acute-phase abortions are likely to be impossible to diagnose from fetal tissues. These results have implications not only for the diagnosis of naturally occurring ovine abortions but also for the understanding of the pathogenesis of toxoplasma-induced abortion.     

Collagen induces cytokine release by fetal platelets: implications in scarless healing

In previous studies the authors demonstrated that unlike adult platelets, fetal platelets respond poorly to collagen. When platelets make contact with the exposed collagen at the site of injury, the result is activation, aggregation, and degranulation with the release of cytokines and growth factors. This sequence of events is well characterized in adult wounds, which heal with an acute inflammatory response and dense scar formation. In sharp contrast, fetal dermal wounds heal without an acute inflammatory response and minimal scar formation. Therefore, the aim of this study was to test the hypothesis that collagen, abundant at the site of dermal injury, is a poor inducer of cytokine release by fetal platelets. This could explain, in part, the minimal inflammation accompanying fetal dermal wound healing. Platelet suspensions from six fetal Yorkshire swine at day 80 of gestation (term, 114 days) were exposed to either arachidonic acid, 0.5 mg/mL, collagen, 0.19 mg/mL, or saline. The release into plasma of transforming growth factor-beta (TGF-beta 1), and platelet-derived growth factor (PDGF)-AB effected by these agents was determined by enzyme-linked immunosorbent assays. Transmission electron microscopy (TEM) was used to correlate the biochemical findings with ultrastructural changes and showed that arachidonate-treated platelets were aggregated and devoid of granules. In contrast, collagen-treated platelets had undergone conformational changes but showed only a moderate change in the quantity and homogeneity of their secretory granules compared with saline-treated controls. There was a significant increase in TGF-beta 1 release into plasma after treatment with collagen (6.64 +/- 0.36 ng/mL) and arachidonate (7.64 +/- 0.77 ng/mL) compared with saline (4.74 +/- 0.36 ng/mL), P < .05. Likewise, PDGF-AB release was significantly higher after collagen (0.22 +/- 0.02 ng/mL) and arachidonate treatment (0.44 +/- 0.04 ng/mL) compared with saline (0.09 +/- 0.02 ng/ mL), P < .05. The authors conclude that fetal platelets actually do release cytokines in response to contact with collagen despite poor aggregation. Therefore, impaired aggregation to collagen cannot solely explain the minimal inflammation after fetal wounding.     

Straightening out the renal tubule: advances in the molecular basis of the inherited tubulopathies

OBJECTIVE: To assess the left ventricular determinants of cardiac output in 12 chronically instrumented fetal lambs, six of which where made anemic. METHODS: Twelve, singleton pregnant ewes were instrumented between 123-126 days gestation and chronic catheters placed in-situ. In six fetal lambs, isovolemic fetal anemia was induced (reducing the hematocrit from a mean of 35% to 20%). In a prospective study, absolute 'beat-to-beat' LV volumes and pressures were obtained (gestational age 131-134 days) using a conductance catheter method and a comparison made with non-anemic fetal lambs. RESULTS: The group of anemic fetuses (n = 6) had a significantly reduced hematocrit as compared to the control group (37% decrease: mean difference--13.4%; P < 0.001). The arterial blood gases of the two groups were not statistically different, with the exception of the pO2 and oxygen content which were significantly lower in the anemic group (P < 0.05). There was no significant change in fetal heart rate, LV preload (as assessed by venous pressure, end-diastolic volume and pressure) or mean arterial pressure between the anemic and control groups. However, a 75% increase in LV stroke volume was observed in the anemic fetal lambs (P < 0.05), secondary to a 61% fall in LV afterload (P < 0.05). There was no significant change in inherent myocardial contractility of the LV, although this did increase to approach statistical significance in the anemic group (P = 0.056). The diastolic time interval was increased by 22% in the anemic fetus, possibly allowing prolonged LV filling time. The indices measuring LV relaxation (Tau and dP/dtmin) were not significantly different in either group. CONCLUSIONS: This is the first study to report absolute left ventricular volumes in the anemic ovine fetus and the relationship of these data to LV pressure during the cardiac cycle. The model used produces a state of moderately severe, non-hydropic, isovolemic, fetal anemia consistent with those previously described. Although the anemic state was not prolonged, an observed increase in LV stroke volume (which is predominantly due to a decrease in afterload) has been described.     

Coordination between glottic adductor muscle and diaphragm EMG activity in fetal lambs in utero

It has previously been reported that active glottic adduction is present during prolonged apneas but absent during periods of breathing movements in fetal lambs in utero. The present study was aimed at examining the precise coordination between fetal breathing movements [diaphragm electromyographic (EMG) activity (Di EMG)] and glottic adduction [thyroarytenoid muscle EMG activity (TA EMG)]. Electrodes for electroencephalogram, eye movements, TA EMG, and Di EMG and an arterial catheter were surgically implanted in fetal lambs 123-142 days postconception. Polygraphic recordings were performed without sedation while the ewe breathed room air (n = 11) or various gas mixtures (hypoxia, n = 5; hyperoxia, n = 4; hypercapnia, n = 5; hypercapnia+hyperoxia, n = 5). Tonic TA EMG was observed throughout >90% of apneas (>6 s) in both non-rapid-eye-movement and rapid-eye-movement sleep, and when Di EMG frequency decreased in rapid-eye-movement sleep. In all but two fetuses, TA EMG was immediately inhibited when Di EMG appeared. Altering blood gases did not modify these results. In conclusion, Di EMG and TA EMG are well coordinated in late gestation in fetal lambs, except in a few cases. These findings may have consequences for understanding the pathogenesis of mixed/obstructive apneas of prematurity.