Regional differences in water content,collagen content,and collagen degradation in the cervix of nonpregnant cows

The cow could be a suitable model for studies concerning functional changes of the cervix. However, as in many species, the bovine cervix becomes softer in texture during the follicular phase of the estrous cycle compared to the luteal phase. In the present study, we explored if changes in the collagen network take place that could be responsible for this phenomenon and if regional differences in water content, collagen content, and collagen degradation along the cross-sectional and longitudinal axes of the cervix were present. Two groups of nonpregnant animals with different progesterone status were studied. One group (n = 11) was under high progesterone influence, and the other group (n = 12) was under low progesterone influence. The water content was derived from the weight of the samples before and after lyophilization. The collagen content and the ratio of collagenous to noncollagenous proteins (hydroxyproline:proline ratio) were determined by performing amino acid analysis on hydrolyzed samples using high-performance liquid chromatography. Collagen denaturation was quantified with a colorimetric assay by determining the amount of hydroxyproline released from samples treated with alpha-chymotrypsine. The water content of the superficial layer of the submucosa was always significantly (P < 0.01) higher than the water content of the deep layer in the vaginal, mid, and uterine segments, but this was unrelated to the progesterone status of the animals. No effect of the tissue layers or of the progesterone status of the animals on the collagen content was observed, but an effect of segment was noted. The collagen content (mug/mg dry wt) in the vaginal segment of the cervix was significantly higher than in the mid (P < 0.05) and the uterine (P < 0.01) segments. The hydroxyproline:proline ratio showed the same pattern as the collagen content. The percentage of collagen denaturation in the superficial layer was always significantly (P < 0.01) higher than that in the deep layer, but no effect of the progesterone status or of the segment along the longitudinal axis was seen. It is concluded that regional differences in collagen biochemistry are present in the cervix of nonpregnant cows, which may account for the difference in firmness of different parts along the circular or the longitudinal axis of the cervix. However, differences in texture of the cervix between the two groups of cows could not be explained by differences in the collagen content, percentage of collagen denaturation, or water content.     

Investigation of total, free, peptide-bound, protein-bound, soluble and insoluble collagen hydroxyproline content in tissues from the Arabian camel (Camelus dromedarius)

This study was conducted to determine the concentration of total, free, peptide-bound, protein-bound, soluble and insoluble collagen hydroxyproline (Hyp) in tissues from the Arabian camel (Camelus dromedarius). Results indicated that there were significant differences in the concentration of total, free, peptide-bound, protein-bound, soluble and insoluble collagen Hyp in various tissues (P < 0.01). Camel kidney showed a significantly high concentration of total, free, peptide-bound and protein-bound Hyp and collagen content as compared to other tissues examined (P < 0.01). Kidney also showed a significantly high concentration of soluble collagen Hyp as compared to other tissues examined (P < 0.01). However, the concentration of insoluble collagen Hyp was significantly high in liver when compared to other tissues (P < 0.01). These variations may result from differences in the collagen structure and/or composition in this species.     

微等离子束对豚鼠皮肤超微结构分析和羟脯氨酸测定

目的观察微等离子束对豚鼠皮肤胶原组织作用效应的组织学和超微结构变化及羟脯氨酸含量测定,探讨微等离子束的作用机理。方法选择15只豚鼠,每只豚鼠背部划分为实验侧和空白对照侧2个等分区域,给予60W/10 kJ微等离子束照射,于作用后即刻、1周后和1月后分别切取实验侧及空白对照部位皮肤行组织病理维多利亚-立春红染色,透射电镜分析和羟脯氨酸检测试剂盒进行含量测定。结果 60 W/10 kJ即刻表现为表皮局灶性出现点阵状改变,部分表皮出现汽化缺失或者坏死变性,真皮浅层胶原组织出现点阵化表现和明显均质化;特殊染色显示微等离子束主要影响真皮胶原纤维,形成局灶性胶原纤维凝集和变性。1周后皮肤浅层胶原组织结构逐渐致密,排列有序,有少量组织细胞。1月后皮肤浅层胶原组织明显增厚,胶原纤维增粗并排列致密,弹力纤维呈局灶性增粗。透射电镜显示微等离子束作用后表皮细胞较完整,细胞间结构正常,但真皮胶原丧失正常结构,细胞结构消失,大量细胞凋亡明显,1月后仍可见少量细胞凋亡的表现但胶原结构逐渐恢复,浅层胶原纤维排列明显致密。羟脯氨酸测定显示微等离子束作用1周后羟脯氨酸含量要高于作用之前,但是差异性不具有统计学意义（P〉0.05）;1月后羟脯氨酸含量要明显高于作用前,差异性具有统计学意义（P〈0.05）。结论微等离子束对豚鼠皮肤胶原组织作用有明显的刺激效应,其主要靶组织为真皮胶原组织,可以明显促进皮肤新生胶原的增生。     

High iNOS mRNA and protein levels during early third trimester suggest a role for NO in prelabor cervical ripening in the bovine

Nitric oxide (NO) plays a key role in the processes leading to cervical softening prior to labor. Inducible nitric oxide synthase (iNOS) contributes most to the increased production of NO during labor, as demonstrated in the rat cervix, or at term pregnancy in women. Changes in expression of iNOS during late gestation have not yet been studied longitudinally in any species, because repeatedly taking biopsies could not be performed. iNOS mRNA (n = 6) and protein expression (n = 3) in serial cervical biopsies of pregnant pluriparous cows taken around days 225, 250, and 275 of pregnancy and within 1.5 hr after calving (d225, d250, d275 and parturition biopsies, respectively) were measured using quantitative RT-PCR and Western blotting. iNOS mRNA expression decreased from the d225 biopsy onwards, differences being significant between the d250 and d275 (P < 0.05) and between the d275 and parturition biopsies (P < 0.05). iNOS protein expression decreased from d225 to d250 onwards. Immunohistochemical analysis of biopsies showed, besides positive staining in endothelium and epithelium, which remained unchanged at different time points, that iNOS expressing cells in the connective tissue cells of early biopsies were predominantly spindle shaped (mostly smooth muscle cells and some fibroblasts). In the parturition biopsies, iNOS reactivity was mainly found in mononuclear leucocytes. These results lead us to suggest that iNOS from spindle shaped cells is involved in prepartum cervical ripening, while iNOS in mononuclear inflammatory cells may be important for local tissue repair mechanisms during postpartum cervical involution.     

Influence of thyroid hormones on collagen content in tissues of guinea pigs.

Collagen fractions content and level of collagen catabolites in body fluids were determined in normal, hypo- and hyperthyroid guinea pigs. An increase of urinary excretion of hydroxyproline and hydroxylysine as well as concentration of these amino acids in blood serum was found in hyperthyroidism, and a decrease was shown in hyperthyroid guinea pigs. Hyperthyroidism stimulated an increase of neutral-salt-soluble and acid-soluble collagen in skin and liver, and a decrease of insoluble collagen in skin as well as increase of all collagen fractions in bone samples. Hypothyroidism induced a decrease of all collagen fractions in skin and liver, and an increase of acid-soluble and insoluble collagen in bone samples.     

Quantitative Evaluation of Collagen Crosslinks and Corresponding Tensile Mechanical Properties in Mouse Cervical Tissue during Normal Pregnancy

The changes in the mechanical integrity of the cervix during pregnancy have implications for a successful delivery. Cervical collagens are known to remodel extensively in mice with progressing gestation leading to a soft cervix at term. During this process, mature crosslinked collagens are hypothesized to be replaced with immature less crosslinked collagens to facilitate cervical softening and ripening. To determine the mechanical role of collagen crosslinks during normal mouse cervical remodeling, tensile load-to-break tests were conducted for the following time points: nonpregnant (NP), gestation day (d) 6, 12, 15, 18 and 24 hr postpartum (PP) of the 19-day gestation period. Immature crosslinks (HLNL and DHLNL) and mature crosslinks (DPD and PYD) were measured using ultra performance liquid chromatography-electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS). There were no significant changes in the total immature crosslink density (HLNL+DHLNL mol per collagen mol) throughout normal mouse gestation (range: 0.31–0.49). Total mature crosslink density (PYD+DPD mol per collagen mol) decreased significantly in early softening from d6 to d15 (d6: 0.17, d12: 0.097, d15: 0.026) and did not decrease with further gestation. The maturity ratio (total mature to total immature crosslinks) significantly decreased in early softening from d6 to d15 (d6: 0.2, d15: 0.074). All of the measured crosslinks correlated significantly with a measure of tissue stiffness and strength, with the exception of the immature crosslink HLNL. This data provides quantitative evidence to support the hypothesis that as mature crosslinked collagens decline, they are replaced by immature collagens to facilitate increased tissue compliance in the early softening period from d6 to d15.     

MMP-2 expression precedes the final ripening process of the bovine cervix

Collagen is denatured in the gradual cervical ripening process during late pregnancy, already before the onset of final cervical ripening at parturition. Matrix Metallo Proteinases (MMPs) might be responsible for this process. To investigate the presence and potential function of MMPs at the different stages of the ripening process, serial cervical biopsies were obtained from 10 cows at Days 185 and 275 of pregnancy (approximately 5 days before calving), at parturition and at 30 days after parturition. The mRNA and protein expression of MMP-1, MMP-2, and MMP-9 and of the tissue inhibitors of MMPs (TIMP)-1 and TIMP-2 were semi-quantitatively determined using RT-PCR, respectively, zymography, Westernblot, and ELISA techniques and the localization of MMP-2 protein and presence of granulocytes by immunohistochemistry and Luna staining. At parturition compared to 185 days pregnancy the MMP-1 protein expression and the numbers of granulocytes were significantly increased by 3 and 26-fold respectively. MMP-2 mRNA and protein expression had already increased 2.5 (P < 0.05) and twofold (P < 0.05) at 5 days before parturition, prior to final ripening. At that time, MMP-2 was present in smooth muscle cells and extra cellular matrix. TIMP-1 mRNA expression was significantly increased at parturition and TIMP-2 mRNA expression peaked at 5 days before parturition. The increased expression of MMP-2 at 5 days before parturition, suggests that in the cow MMP-2 is responsible for collagen denaturation in the last part of gradual cervical ripening, while MMP-1 and MMP-9 are only active during the final cervical ripening process at parturition.     

Cervical diameter in relation to uterine and cervical EMG activity in early postpartum dairy cows with retained placentas after PGF2alpha induced calving

The cervix must regain its normal diameter after parturition. Until now, little has been known about the pattern of cervical closure and the possible influences of myometrial and cervical contractions in this process. We continuously measured the cervical diameter with ultrasound cervimetry during the first 48h after calving in six cows with retained fetal membranes, while uterine (n=6) and cervical outer muscular layer (n=4) electromyographic (EMG) activity was measured with bipolar EMG electrodes. We found that the cervical diameter which was 6.2cm (+/-0.7) at 1.4h after calving, initially increased to 9.0cm (+/-1.0) during the first 14.8h (+/-2.8) postpartum. After this time, the diameter decreased gradually to 5.3cm (+/-1.0) at 48h after calving. The overall EMG activity after parturition decreased by 59% (+/-6) and 35% (+/-17) for the uterus and cervix, respectively. The decrease in EMG activity was due to a 50% (+/-7) decrease in EMG amplitudes of the myometrium; the EMG amplitudes of the cervix decreased by only 8% (+/-21) (P>0.05). At the same time in the cervix, burst frequency decreased by 69% (+/-17), while the decrease in burst frequency of the myometrium was only 11% (+/-5) (P>0.05). Uterine myometrial and cervical EMG activity after parturition showed burst patterns. These contractions of the uterus and cervix were accompanied by and correlated with transient dilatations of the caudal cervix. This could have functional relevance in the evacuation of the uterus.     

Comparative analysis of the structure and thermal stability of sea urchin peristome and rat tail tendon collagen

We have purified collagen from two distinct sources; the vertebrate, rat tail tendon and an invertebrate, sea urchin adult tissue, the peristome. The collagenous nature of the purification products was confirmed by amino acid compositional analysis. Both preparations had high contents of glycine and proline residues and hydroxyproline was also present. The total pyrrolidine (proline+hydroxyproline) content decreased from 17.9 mole% in rat tail collagen to 12.9 mole% in peristome collagen. Distinctly different circular dichroic spectra were measured for these collagens. Analyses of spectra, measured as a function of temperature, revealed distinct thermal denaturation profiles. The melting temperature for rat tail collagen was 38.5 degrees C, while the corresponding value for peristome collagen was significantly lower at 27 degrees C. A similar thermal denaturation profile was obtained for rat tail collagen in digestion experiments using a 41-kDa gelatinase activity, isolated from sea urchin eggs. These results identify structural differences between a typical, vertebrate type I fibrillar collagen and an echinoderm collagen which serves as a constituent of a mutable connective tissue. These differences may relate to the functional roles played by collagen in these distinctly different tissues.     

Effects of training and anabolic steroids on collagen synthesis in dog heart.

The effects of endurance training and anabolic steroid (Methandienone 1.5 mg.kg-1 p. o. daily) and their combination on regional collagen biosynthesis and concentration in the hearts of male beagle dogs were studied by measuring prolyl 4-hydroxylase (PH) activity and hydroxyproline (HYP) concentration. The PH (P less than 0.05) and HYP (P less than 0.05) were both greater in the subendocardinal layer than in the subepicardium (EPI) of the left ventricular wall in controls, whereas opposite gradients (P less than 0.05) were observed in the right ventricle. Endurance exercise caused an increase of PH activity in EPI of the left ventricular wall (P less than 0.01). The HYP concentration increased in both layers of the right ventricle in the exercise plus steroid group (P less than 0.05). The results suggest that transmural differences exist in the rate of collagen synthesis and concentration in canine cardiac ventricles and that endurance exercise may accelerate collagen synthesis in EPI of the left ventricle and the combination of exercise and anabolic steroid causes an increase in collagen concentration in the right ventricular wall.     

Cervical changes in estrogen receptor alpha,oxytocin receptor,LH receptor,and cyclooxygenase-2 depending on the histologic compartment,longitudinal axis,and day of the ovine estrous cycle

The aim was to investigate the histologic distribution of estrogen receptor α (ERα), oxytocin receptor (OxR), LH receptor (LHR), and cyclooxygenase-2 (COX-2) in the cervix of the ewe during the estrous cycle. Immunohistochemistry was performed in the cranial and caudal cervix of Corriedale ewes on Day 1 (n = 6), 6 (n = 5), or 13 (n = 6) after estrous detection (Day 0). The ERα proportional score (%ERα nuclei) was lower in the cranial cervix than in the caudal cervix, whereas the OxR and COX-2 immunostaining areas (%areas) were greater in the cranial cervix than in the caudal cervix (P < 0.04). The %ERα nuclei decreased from Days 1 to 13 in luminal epithelia, but increased from Days 1 to 6 or remained unchanged in stromata (P < 0.003). The %OxR area was higher on Day 6 than on Days 1 and 13 in the superficial glandular epithelium, and increased from Days 1 to 13 in the deep glandular epithelium (P < 0.04). The %LHR area increased during the estrous cycle in luminal epithelia and fold stroma (P < 0.004). The %COX-2 area was restricted to epithelia, and it was lower on Day 1 than on Days 6 and 13 in luminal epithelia (P < 0.05). Differences in ERα, OxR, LHR, and COX-2 between cranial and caudal cervical zones indicate different physiological functions, and their cyclic variations in the cervical epithelia, in contrast to little or no variations in the stroma, suggest a hormone-responsive driving role of epithelia in cervical function.     

Synthesis and turnover of collagen precursors in rabbit skin

1. The rate of synthesis of [(14)C]hydroxyproline by rabbit skin was studied in vitro and in vivo. 2. The soluble collagen fractions were shown to have a very rapid turnover. The 0.15m-sodium chloride-extractable collagen showed t((1/2)) values of 1.2hr. in vitro and 12hr. in vivo. The 0.5m-sodium chloride-extractable collagen exhibited a t((1/2)) value of 20hr. in vivo. 3. Under the conditions used it was not possible to obtain radioactive insoluble collagen in vitro. 4. A significant amount of soluble collagen is lost before it becomes insoluble. 5. These observations may help to explain why large amounts of peptide-bound hydroxyproline appear in the urine during periods of rapid collagen synthesis.     

Skeletal muscle collagen content in humans after high-force eccentric contractions.

The purpose of this study was to investigate the effects of high-force eccentric muscle contractions on collagen remodeling and on circulating levels of matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) in humans. Nine volunteers [5 men and 4 women, mean age 23 (SD 4) yr] each performed a bout of 100 maximum voluntary eccentric contractions of the knee extensors. Muscle biopsies were taken before exercise and on days 4 and 22 afterward. Image analysis of stained tissue sections was used to quantify endomysial collagen staining intensity. Maximum voluntary contractile isometric force was recorded preexercise and on days 1, 2, 3, 4, 8, 11, and 14 postexercise. Venipuncture blood samples were also drawn on these days for measurement of serum creatine kinase activity and concentrations of MMP-9, TIMP-1, TIMP-2, and the MMP-2/TIMP-2 complex. Maximum voluntary contractile force declined by 39 +/- 23% (mean +/- SD) on day 2 postexercise and recovered thereafter. Serum creatine kinase activity peaked on day 4 postexercise (P < 0.01). Collagen type IV staining intensity increased significantly on day 22 postexercise to 126 +/- 29% (mean +/- SD) of preexercise values (P < 0.05). Serum MMP-9 levels increased on day 8 postexercise (P < 0.01), and serum TIMP-1 was also significantly elevated on days 1, 2, 3, 4, and 14 postexercise (P < 0.05). These results suggest that a single bout of eccentric muscle contractions results in remodeling of endomysial type IV collagen, possibly via the MMP pathway.     

Proinflammatory cytokines found in amniotic fluid induce collagen remodeling, apoptosis, and biophysical weakening of cultured human fetal membranes

The mechanisms by which fetal membranes (FM) rupture during the birth process are unknown. We have recently reported that FM weaken, at least in part, because of a developmental process of extracellular matrix remodeling and apoptosis. We now hypothesize that cytokines that normally increase in amniotic fluid at term induce FM collagen remodeling and apoptosis with concomitant weakening. Full-thickness FM fragments were cultured with (0-100 ng/ml) or without tumor necrosis factor (TNF) or interleukin 1, beta (IL1B). Physical properties were then examined with specially adapted industrial rupture strength testing equipment. Cultured FM were also evaluated for evidence of collagen remodeling and apoptosis. Cytokine-treated FM exhibited a dose-dependent decrease in strength and work to rupture. Compared with controls, the highest TNF dose caused maximal decrease in FM rupture strength (13.2 +/- 1.2 N versus 3.8 +/- 1.5 N; P = 0.0003) and work to rupture (0.035 +/- 0.005 J versus 0.005 +/- 0.002 J; P < 0.0001). The highest IL1B dose also decreased rupture strength (12.9 +/- 3.2 versus 4.6 +/- 1.1 N; P = 0.0027) and work to rupture (0.018 +/- 0.005 J versus 0.005 +/- 0.002 J; P < 0.01). Matrix metalloproteinase 9 (MMP9) protein increased, tissue inhibitor of matrix metalloproteinase 3 (TIMP3) protein decreased, and poly (ADP-ribose) polymerase (PARP1) cleavage increased with increasing TNF or IL1B doses (all P < 0.05), suggesting collagen remodeling and apoptosis. TNF and IL1B cause significant weakening of cultured FM. Both cytokines induce biochemical markers in the FM in a manner characteristic of the weak zone of FM overlying the cervix. TNF and or IL1B may be involved in the development of the weak zone of the FM.     

Changes in the histomorphology of the canine cervix through the oestrous cycle

The importance of canine reproduction is steadily increasing and little is known about the canine cervix so far. The aim of this study was to describe the histomorphology of the canine cervix and to determine its correlation to the stage of oestrous cycle and to circulating concentrations of progesterone (P4) and estradiol-17beta (E2), respectively. Ovariohysterectomy (OHE) was performed in 33 non-pregnant, healthy, intact bitches in defined stages of oestrous cycle (proestrus N = 5, estrus N = 7, early diestrus N = 5, late diestrus N = 11, anestrus N = 5). The entire cervix was collected for histological evaluation of the epithelial layer (number of layers, thickness), the stroma (number of layers, thickness, density, structure, and distribution of elastic fibres) and the average area and density of cervical glands as well as blood vessels. These parameters were evaluated in all the three parts of the cervix (cranial, middle, and caudal or vaginal part). The cervix showed the typical structure with a superficial epithelium, a lamina propria with cervical glands and vascular structures and a tunica muscularis below. Folds in the superficial epithelium were only observed in 54% females (N = 18). Epithelial thickness (P < 0.0001), number of glands (P < 0.05), mean area of glands (P < 0.0001), mean area of venous vessels (P < 0.0001), number of arterial vessels (P = 0.02), number of mast cells and number of eosinophilic granulocytes per mm² (P < 0.01) were significantly influenced by the stage of cycle. The following factors were significantly influenced by the localisation: number of epithelial layers (P < 0.0001), thickness of stroma (P < 0.0001) and mean number of glands (P < 0.01). Only mean area of venous vessels was significantly influenced by the stage of cycle and the localisation (P < 0.01). Besides, P4 was significantly correlated to number of glands per mm² (P < 0.0001), mean area of venous vessels (P < 000.1) and number of mast cells (P < 0.01).This study provided detailed information about the histomorphological structure of the cervix in non-pregnant bitches and showed that the cervix undergoes cyclic changes during the canine oestrous cycle, in particular in association with circulating progesterone concentration.     

Collagenase activity in cervical tissue of the non-pregnant and pregnant human cervix

Collagen degradation has been suggested to play an important role in the process of cervical ripening at term. After giving informed consent tissue samples were taken from the posterior lip of 10 non-pregnant women of child-bearing age, 10 women having a termination of pregnancy at 9-12 weeks' gestation and 12 women who had had normal spontaneous deliveries at term. Collagenase activity was measured by a specific technique using native, triple helical collagen. Proteolytic activity was determined with commercially available 14C-methaemoglobin as substrate. For identification of collagen fragments SDS-polyacrylamid-gel-electrophoresis was performed using a modified Laemmli-technique. Collagenase and protease activity were found in all extracts of the different groups. There was a marked increase in enzymatic activities in the postpartum tissue samples suggesting that significant collagen breakdown had taken place. The absence of typical collagen fragments in the SDS-electrophoresis seemed to be due to a very rapid further degradation of the fragments by proteases into small peptides which then readily left the cervix. The present findings confirm the essential role of proteolytic enzymes in cervical dilatation at term.     

EMG activity of the muscular and stromal layer of the cervix in relation to EMG activity of the myometrium and cervical dilatation in PGF2alpha induced parturition in the cow

The goal of this study was to quantify and characterize the electromyographic (EMG) activities in the cervical outer muscular layer and in the cervical stromal layer, and to characterize their relationship with myometrial EMG activity and cervical dilatation during PGF2alpha-induced parturition in term pregnant cows. We continuously measured the EMG activity of the uterine myometrium and cervical outer muscular layer as well as the cervical stromal layer in five cows using bipolar electrodes while at the same time measuring changes in the cervical diameter with ultrasound cervimetry. This we did from the moment a prostaglandin analogue was injected until the expulsion of the calf. In contrast to the cervical stromal layer, the cervical outer muscular layer showed distinct EMG activity, which began to increase at about the same time as the EMG activity of the myometrium, i.e. some 12 h before the start of cervical dilatation. However, the rate of this increase was lower than in the myometrium and it was not characterized, like in the myometrium, by an increase in maximum EMG amplitude. Although the cervical outer muscular layer showed contracture and contraction like EMG activity in unison with in the myometrium, it was also characterized by a more irregular EMG activity, which occurred independently from the myometrium. These data suggest that while the outer muscular layer of the cervix may be considered to be a caudal continuation of the myometrium, it also displays activity independently from the myometrium. The physiological relevance of this activity remains to be explored.     

Contraction of fibroblast-containing collagen gels: Initial collagen concentration regulates the degree of contraction and cell survival

Remodeling of extracellular matrix involves a number of steps including the recruitment, accumulation, and eventual apoptosis of parenchymal cells as well as the production, organization, and rearrangement of extracellular matrix produced by these cells. The culture of fibroblasts in three-dimensional gels made of type I collagen has been used as a model of tissue contraction which characterizes both wound repair and fibrosis. The current study was designed to determine the effect of initial collagen concentration on the ability of fibroblasts to contract collagen gels and on cell survival. Native type I collagen was extracted from rat tail tendons and used to prepare collagen gels with varying collagen concentrations (0.75-2.0 mg/ml). Human lung fibroblasts (HFL-1) were cast into the gels and cultured in Dulbecco modified Eagle medium with 0.1% fetal calf serum for 2 wk. The gel size, collagen content, and deoxyribonucleic acid (DNA) content were determined. Gels prepared with an initial concentration of 0.75 mg/ml contracted more rapidly and to a smaller final size than gels prepared from 2 mg/ml initial collagen concentration (final size 7.1 versus 36.4% of initial size, P < 0.01). There was no significant degradation of the collagen in the gels under either condition. Hence, the dramatically increased contraction of the lower density gels resulted in a higher final density (P < 0.01). Cell density was estimated from DNA content. In low initial density gels, the final DNA content was significantly less than that in higher initial density gels (0.73 versus 1.88 microg/gel, P < 0.05). This was accompanied by an increased percentage of apoptotic cells at day 14 (43.3 versus 34.1%, P < 0.05). If the gels were maintained in the attached state which largely prevents contraction, apoptosis was significantly reduced, suggesting that contraction rather than matrix composition was a requirement for the increased apoptosis. In summary, these findings indicate that the initial matrix composition can lead to differing outcomes during fibroblast-mediated wound contraction.     

Relationship between dilatation of the rat uterine cervix and a small dermatan sulfate proteoglycan

Cervical linear circumference (lo), extensibility and rate of creep, and the content and concentration of collagen and proteoglycans were determined on uterine cervices of rats at different reproductive stages. The inner circumference increased from 9 +/- 3 (SD) mm at the nongravid stage to a maximum of 41 +/- 5 mm at term; a significant drop to 23 +/- 2 mm occurred by 4 h postpartum with a further drop to 18 +/- 4 mm by 1 day postpartum. The extensibility and rate of creep reached their maxima 1 day before term and returned to the nongravid value by 1 day postpartum. The small (Mr = 95,000) type II dermatan sulfate proteoglycan, the major cervical proteoglycan, increased from 43 +/- 6 micrograms per cervix at the nongravid stage to 196 +/- 33 micrograms at term. The amount of this proteoglycan decreased significantly by 35% to 126 +/- 5 micrograms within 4 h postpartum and declined further to 79 +/- 16 micrograms by 1 day postpartum. The total cervical collagen content increased less than 2-fold during pregnancy, from 3.5 +/- 0.5 to 6.3 +/- 0.7 mg; a decline to 5.8 mg by 1 day postpartum was not significant. The ratio of small proteoglycan: collagen increased 2.5-fold between the nongravid state and term, then returned to the nongravid value by 1 day postpartum. Significant correlations were found between the lo and the amount of small proteoglycan per cervix (r = 0.86; n = 69) and between lo and the ratio of small proteoglycan:collagen (r = 0.83; n = 50) when data from every reproductive stage were combined. A mechanism is proposed whereby the interaction of the proteoglycan with collagen fibers might alter mechanical properties and contribute to cervical dilatation and its rapid reversal.     

Cervical dilatation related to uterine electromyographic activity and endocrinological changes during prostaglandin F(2alpha)-induced parturition in cows

The temporal relationship between changes in cervical dilatation, uterine electromyographic (EMG) activity, and maternal plasma concentrations of estradiol 17beta (E(2)), progesterone (P(4)), and 13,14-dihydro-15-keto-prostaglandin-F(2alpha) (PGFM), was investigated in six parturient cows. Calving was induced with a single injection of a synthetic analogue of prostaglandin F(2alpha) (PG) on Day 274 of gestation. Cervical dilatation was measured continuously by measuring the transit time between two implanted ultrasound crystals while at the same time uterine EMG activity was measured through two silver electrodes sutured on the myometrial surface until the expulsive stage of calving had been reached. In blood samples collected at 4-h intervals, starting at the moment of PG injection, the mean plasma E(2) concentration gradually increased and was significantly elevated at 28 h after PG injection. At 4 h after PG treatment, the mean P(4) concentration had dropped significantly and continued to decrease until a value of around 1 ng/ml was reached, where it stayed until the onset of expulsion. Mean plasma PGFM concentrations increased steadily after PG injection, reaching significantly elevated concentrations at 20 h after treatment. In the five cows that delivered calves in anterior positions, uterine EMG activity, expressed as root mean square (RMS in microV), started to increase at a mean interval (+/- SD) of 13.1 +/- 3.7 h following PG treatment. The increase in EMG activity was significantly correlated with changes in plasma PGFM concentrations. In these cows, dilatation of the caudal cervix started after a mean (+/- SD) interval of 28.5 +/- 1.5 h following PG treatment and dilatation progressed at a mean (+/- SD) rate of 2.25 +/- 0.24 cm/h. In one cow with a calf in the posterior position, uterine EMG activity and dilatation started at 15.8 h and 31.8 h, respectively, after induction of calving. We conclude that a predictable sequence of physiological changes occurs around induction of calving, which allows specific timing of future studies on cellular and biochemical changes within the cervix during parturition.