Uterine Vascular Perfusion Research Articles

Abstract P235: The Absence Of Inhibitory G Protein Regulation By Rgs2 Increases Basal Phosphodiesterase Activity To Decrease Forskolin-stimulated Camp Generation And Enhances Myogenic Response In Uterine Arteries Of Non-pregnant Mice

G protein regulation by regulators of G protein signaling (RGS) proteins is crucial to maintaining normal vascular tone for promoting uterine vascular bed perfusion. However, the specific regulatory mechanisms involved are incompletely understood. Previously, we showed that loss of regulation of Gi/o and Gq/11 by RGS2 contributes to enhanced myogenic tone and decreased uterine blood flow in non-pregnant mice. Similarly, RGS5, closely related to RGS2, was found to fine-tune G protein signaling and modulate GPCR-evoked vascular reactivity. RGS2 and RGS5 are co-expressed in the uterine vascular bed. However, it is unclear whether their regulatory activities are coordinated in fine-tuning G protein signaling to modulate vascular tone. Here, we tested whether the combined activity of RGS2 and 5 reduces myogenic tone by negatively regulating Gi/o signaling to promote cAMP-dependent uterine artery relaxation. We used commercially available kits to measure forskolin-stimulated cAMP generation and basal phosphodiesterase (PDE) activity in freshly isolated uterine arteries from non-pregnant wild type, Rgs5 KO and Rgs2 KO mice. Basal tone and myogenic response to increasing intraluminal pressure were assessed using ex vivo pressure myography. We found that the absence of RGS2 but not RGS5 decreased forskolin-stimulated cAMP generation. Conversely, basal PDE activity was augmented in both Rgs2 KO and Rgs5 KO arteries, though the increase was less robust in Rgs5 KO arteries. Basal tone and myogenic response were reduced in Rgs5 KO arteries compared to WT or Rgs2 KO arteries in the absence or presence of nitric oxide synthase blockade with L-NAME. Activating Gi/o with dopamine increased myogenic constriction in all genotypes but less robustly in Rgs5 KO arteries, while pre-incubation of the vessels with exogenous cAMP or PDE inhibition with IBMX normalized maximal myogenic response to similar levels in all groups. Collectively, these findings indicate that altered Gi/o regulation due to RGS2 absence enhances myogenic tone by promoting PDE-mediated inhibition of cAMP-dependent vasodilation. While both RGS2 and RGS5 negatively regulate this novel Gi/o-PDE-cAMP signaling pathway, RGS5 dampens the inhibitory efficacy of RGS2 towards Gi/o in uterine arteries.

Exogenous L-arginine administration improves uterine vascular perfusion, uteroplacental thickness, steroid concentrations and nitric oxide levels in pregnant buffaloes under subtropical conditions.

Heat stress (HS) during pregnancy adversely affects uterine vascular perfusion and foetal development. L-arginine (L-Arg), a nitric oxide (NO) precursor, has been proven to enhance an organ's vascular perfusion. Therefore, this study aimed to examine the effect of L-Arg administration on the pregnant buffaloes' uterine haemodynamics and uteroplacental thickness under environmental HS conditions. For this purpose, pluriparous (n=12) HS-pregnant buffaloes (mid-gestation, 180-190 days; temperature humidity index >85) were haphazardly assigned into two groups, either administered a single intravenous (IV) bolus of 5mg/kg BW of L-Arg Hcl (n=6; ARG) or received normal saline (0.9%; IV; 25 ml) that served as a control group (n=6; CON). Combined uteroplacental thickness (CUPT, mm), middle uterine arteries' (MUA) diameter (mm) and uterine Doppler haemodynamics (ipsilateral and contralateral to the pregnant horn) were examined utilizing B-mode and colour Doppler ultrasonography, respectively. The serum levels of progesterone (P4), estradiol 17β (E2), nitric oxide (NO) and total antioxidant capacity (TAC) were assayed using commercial kits. The experimental time points were - 1, 0, 2, 4, 24, 48, 72, 96 and 120 h post-L-Arg administration. The ARG group showed higher (p < .05) CUPT and MUA diameter values starting at 24 h post-treatment and onward than the control buffalo cows. In addition, there were improvements (p < .05) in the blood flow parameters in the ipsilateral MUA after L-Arg treatment evidenced by lower values of pulsatility and resistive indices starting at 48 and 4h, respectively and onward, and higher values of peak systolic velocity, coloured areas towards the pregnant uterine horn (both 24-72 h). Increases in the NO levels were found during the period between 4 and 120 h in the ARG group compared to the CON group. Moreover, significant increases in the E2 and P4 means were noted in the ARG group, especially at 24 h onward, respectively compared to the CON group. Concerning the TAC status, neither the administration nor the hours affected serum TAC levels in the HS pregnant buffaloes (CON and ARG). In conclusion, L-Arg administration improved uteroplacental thickness and enhanced uterine haemodynamics, NO levels, and steroids production in mid-pregnant buffalo cows under environmental heat stress conditions which could improve foetal growth and development.

The efficiency of intrauterine infusion of platelet-rich plasma in the treatment of acute endometritis as assessed by endoscopic, Doppler, oxidative, immunohistochemical, and gene expression alterations in jennies

This study used autologous platelet-rich plasma (PRP) to treat acute endometritis in jennies with follow-up for alterations in uterine hemodynamics, endoscopic, immunohistochemistry, oxidant/antioxidant imbalance, pro-inflammatory regulatory molecules, and transmembrane mucin expressions. Ten jennies suffering from endometritis (acute type; n = 10) were included in the study. PRP was prepared from each animal and two intrauterine infusions one week apart were administrated. Examination and follow-up were done physically, ultrasonographically, endoscopically and samples were taken for histopathology, immunohistochemistry, and bacteriological examination. Blood and uterine fluid samples were taken to estimate biochemical and oxidative stress alterations. Expression of TRAF6 and MUC1 genes was investigated in uterine fluid, at days −1 (day of diagnosis establishment), 7, 14, and 21. Uterine bacteriological examination showed a decrease in bacterial isolates after PRP treatment. The uterine thickness and uterine vascular perfusion as illustrated by color Doppler ultrasonography were significantly decreased in jennies treated by PRP. Uterine spectral wave pattern showed a significant linear increase in pulsatility index only. Three weeks after first PRP treatment, white light endoscopic examination revealed normal uterine body mucosa and uterine horn folds. A high nuclear factor (NF-κB) expression was seen in the mononuclear cells. A significant reduction in oxidative stress biomarkers in both serum and uterine fluid was recorded after PRP treatment. The TRAF-1 gene expression significantly decreased gradually after intrauterine PRP infusion. The MUC-1 gene expression significantly decreased gradually after intrauterine PRP infusion. Both genes were within normal levels by week 3. Endometritis in jennies is associated with an oxidative process, alterations in serum biochemical parameters, Doppler indices, endoscopic appearance, high NF-κB expression, and upregulation of TRAF-1 and MUC-1 expressions. Two intrauterine infusions of autologous PRP restored normal endometrial appearance after acute endometritis.

Melatonin and CIDR improved the follicular and luteal haemodynamics, uterine and ovarian arteries vascular perfusion, ovarian hormones and nitric oxide in cyclic cows.

This study hypothesizes that melatonin with exogenous progesterone (CIDR) can improve follicular, luteal, ovarian and uterine haemodynamic of heat-stressed cows. Holstein cows (N=12) studied for two spontaneous oestrous cycles during winter then divided equally during summer into the CIDR group received CIDR for 7days and the melatonin group (Mel) received three injections of melatonin (75mg/head) at the CIDR insertion, removal and ovulation days. Blood samples were collected to assay oestradiol (E2), progesterone (P4) and nitric oxide (NO). On day 0 (Ovulation), Mel had more small follicles (p<.05), higher ipsilateral and contralateral ovarian arteries (Ov.A.) peak systolic velocity (PSV), higher ipsilateral uterine artery (Ut.A.) PSV (p=.031) and blood flow volume (BFV), also Mel elevated contralateral Ut.A. PSV and BFV (p<.0001) but lowered contra Ut.A. pulsatility index (PI, p<.0001), E2 (p<.01) and NO (p<.0001). Mel increased the corpus luteum diameter (CL, p<.001), coloured area (p<.007) and P4 (p<.0001) on day 5 and reduced them (p<.05; p<.01) on Day 14. On day 10, Mel obtained CL diameter (p<.03) and coloured area (p<.002) of spontaneous that was higher than CIDR and decreased P4 (p<.003). Mel increased CL diameter, area and coloured area and decreased them thereafter. Mel increased the ipsilateral ovarian and uterine arteries PSV and BFV before ovulation and until day 8. Mel increased P4 and decreased NO until days 6 and 14. In conclusion, the improvement in follicular, luteal, ovarian and uterine haemodynamic and the decrease of NO production proved our hypothesis Melatonin doses higher than 75mg/head is recommended to improve the heat-stressed cow's fertility.

Utility of indocyanine green injection in patients with cervical cancer besides the identification of sentinel lymph node (Review).

Due to the wide introduction of screening tests, patients with neoplastic diseases of the uterine cervix trend to be diagnosed in early stages of the disease, and less invasive surgical procedures are needed in certain cases. In this respect, the technique of sentinel lymph node dissection has been widely implemented as part of the therapeutic strategy, indocyanine green (ICG) being one of the most reliable markers for sentinel lymph node detection. However, it seems that this agent is extremely useful in order to achieve better short-term and long-term results after cervical cancer surgery, due to its capacity to determine the uterine vascular perfusion in cases in which conservative treatment, such as trachelectomy, is performed, as well as to prevent and treat lower limb lymphedema. A literature review was conducted of the studies which focused on the role of ICG utility in cervical cancer patients besides sentinel lymph node detection, special focus was given to vascularization preservation and lower limb lymphedema prevention and treatment.

Pregnancy rates after manual puncture of equine embryos following direct transfer or vitrification

In order to determine whether differences in uterine blood flow between pregnant and non-pregnant mares can be used to predict the presence of the equine embryo prior to flushing in an embryo transfer program, power Doppler ultrasonography was used on a total of 52 mares on days 7 or 8 post-ovulation. Computer analysis of Doppler images was subsequently performed using ImageJ v1.48 software. Vascular perfusion of the endometrium was analyzed using spot meter techniques, measuring mean pixel intensity and area of blood flow. Mares with positive flushings presented a higher uterine blood flow area (one embryo: 54.01 ± 2.27 mm2 or two embryos: 61.01 ± 6.73 mm2) prior to embryo recovery compared to barren mares (21.77 ± 2.22 mm2) (p ≤ 0.05). However, significant differences in vascular perfusion were not detected between single or twin pregnancies. Blood flow area appears to be a good predictor for differentiation between pregnant and non-pregnant mares with an AUC: 0.869; p ≤ 0.001 and an optimal cut-off value of 37.21 mm2. Both the mare’s age and day of embryo recovery caused effects on uterine vascular perfusion. According to Youden’s J statistics the uterine blood flow area of young pregnant mares was greater than 25.4 mm2 on day 7 (with a sensitivity of 75% and a specificity of 87.5%) and greater than 21.02 mm2 on day 8 post-ovulation (with a sensitivity of 93.8% and a specificity of 100%). The uterine blood flow area in adult pregnant mares was greater than 41.4 mm2 on day 7 (with a sensitivity of 80% and a specificity of 85.5%) and greater than 35.55 mm2 on day 8 after ovulation (with a sensitivity of 97.2% and a specificity of 85.7%). Evaluation on day 8 is therefore considered to be more reliable. Older and middle aged pregnant mares (5–18 years old) had increased uterine vascularization compared to young pregnant mares (2–5 years old) (p ≤ 0.001). Conversely, older barren mares showed higher endometrial vascularity (35.06 ± 2.56 mm2) than young (17.21 ± 1.26 mm2) and middle aged non-pregnant mares (23.84 ± 1.50 mm2) (p ≤ 0.05). We hypothesized that the higher blood flow area seen in older barren mares may be a consequence of a subclinical endometritis due to repeated flushing for embryo recovery. The results of the present study indicate that power Doppler ultrasound combined with computer assisted analysis of images are reliable techniques to detect early pregnancy prior to embryo recovery.

Effect of season on pregnancy rates after embryo transfer in cyclic, synchronized and selected horse recipients

In order to determine whether differences in uterine blood flow between pregnant and non-pregnant mares can be used to predict the presence of the equine embryo prior to flushing in an embryo transfer program, power Doppler ultrasonography was used on a total of 52 mares on days 7 or 8 post-ovulation. Computer analysis of Doppler images was subsequently performed using ImageJ v1.48 software. Vascular perfusion of the endometrium was analyzed using spot meter techniques, measuring mean pixel intensity and area of blood flow. Mares with positive flushings presented a higher uterine blood flow area (one embryo: 54.01 ± 2.27 mm2 or two embryos: 61.01 ± 6.73 mm2) prior to embryo recovery compared to barren mares (21.77 ± 2.22 mm2) (p ≤ 0.05). However, significant differences in vascular perfusion were not detected between single or twin pregnancies. Blood flow area appears to be a good predictor for differentiation between pregnant and non-pregnant mares with an AUC: 0.869; p ≤ 0.001 and an optimal cut-off value of 37.21 mm2. Both the mare’s age and day of embryo recovery caused effects on uterine vascular perfusion. According to Youden’s J statistics the uterine blood flow area of young pregnant mares was greater than 25.4 mm2 on day 7 (with a sensitivity of 75% and a specificity of 87.5%) and greater than 21.02 mm2 on day 8 post-ovulation (with a sensitivity of 93.8% and a specificity of 100%). The uterine blood flow area in adult pregnant mares was greater than 41.4 mm2 on day 7 (with a sensitivity of 80% and a specificity of 85.5%) and greater than 35.55 mm2 on day 8 after ovulation (with a sensitivity of 97.2% and a specificity of 85.7%). Evaluation on day 8 is therefore considered to be more reliable. Older and middle aged pregnant mares (5–18 years old) had increased uterine vascularization compared to young pregnant mares (2–5 years old) (p ≤ 0.001). Conversely, older barren mares showed higher endometrial vascularity (35.06 ± 2.56 mm2) than young (17.21 ± 1.26 mm2) and middle aged non-pregnant mares (23.84 ± 1.50 mm2) (p ≤ 0.05). We hypothesized that the higher blood flow area seen in older barren mares may be a consequence of a subclinical endometritis due to repeated flushing for embryo recovery. The results of the present study indicate that power Doppler ultrasound combined with computer assisted analysis of images are reliable techniques to detect early pregnancy prior to embryo recovery.

Power Doppler can detect the presence of 7–8 day conceptuses prior to flushing in an equine embryo transfer program

In order to determine whether differences in uterine blood flow between pregnant and non-pregnant mares can be used to predict the presence of the equine embryo prior to flushing in an embryo transfer program, power Doppler ultrasonography was used on a total of 52 mares on days 7 or 8 post-ovulation. Computer analysis of Doppler images was subsequently performed using ImageJ v1.48 software. Vascular perfusion of the endometrium was analyzed using spot meter techniques, measuring mean pixel intensity and area of blood flow. Mares with positive flushings presented a higher uterine blood flow area (one embryo: 54.01 ± 2.27 mm2 or two embryos: 61.01 ± 6.73 mm2) prior to embryo recovery compared to barren mares (21.77 ± 2.22 mm2) (p≤0.05). However, significant differences in vascular perfusion were not detected between single or twin pregnancies. Blood flow area appears to be a good predictor for differentiation between pregnant and non-pregnant mares with an AUC: 0.869; p≤0.001 and an optimal cut-off value of 37.21 mm2. Both the mare’s age and day of embryo recovery caused effects on uterine vascular perfusion. According to Youden’s J statistics the uterine blood flow area of young pregnant mares was greater than 25.4 mm2 on day 7 (with a sensitivity of 75% and a specificity of 87.5%) and greater than 21.02 mm2 on day 8 post-ovulation (with a sensitivity of 93.8% and a specificity of 100%). The uterine blood flow area in adult pregnant mares was greater than 41.4 mm2 on day 7 (with a sensitivity of 80% and a specificity of 85.5%) and greater than 35.55 mm2 on day 8 after ovulation (with a sensitivity of 97.2% and a specificity of 85.7%). Evaluation on day 8 is therefore considered to be more reliable. Older and middle aged pregnant mares (5–18 years old) had increased uterine vascularization compared to young pregnant mares (2–5 years old) (p≤0.001). Conversely, older barren mares showed higher endometrial vascularity (35.06 ± 2.56 mm2) than young (17.21 ± 1.26 mm2) and middle aged non-pregnant mares (23.84 ± 1.50 mm2) (p≤0.05). We hypothesized that the higher blood flow area seen in older barren mares may be a consequence of a subclinical endometritis due to repeated flushing for embryo recovery. The results of the present study indicate that power Doppler ultrasound combined with computer assisted analysis of images are reliable techniques to detect early pregnancy prior to embryo recovery.

Uterine Vascular Perfusion and Involution During the Postpartum Period in Mares

In horses, limited data is found regarding the vascular events during uterine involution at the puerperal period. Thus, we aimed to evaluate the morphological aspects (size of uterus and intrauterine fluid content) and the hemodynamics (endometrial and mesometrial vascular perfusion) of the uterus during its postpartum involution process. Ten mares were daily scanned by transrectal color Doppler ultrasonography from the first day postpartum (d1) to the 16th day after first postpartum ovulation (D0 = ovulation). The formerly gravid horn (GH) and formerly nongravid horn (NH) were individually evaluated. A reduction (P < .05) in the uterine diameter was observed during the first 7 days postpartum, and the rate of uterine involution decreased after this period. The involution was completed on d21 and d24 for the NH and GH, respectively. Presence of intrauterine fluid was present in large amounts between d1 and d2 postpartum, followed by a decrease (P < .05) between d4 and d7. No fluid was observed after d16 postpartum or after the third day postovulation (D3). During the early postpartum period, an increase (P < .05) in the endometrial and mesometrial vascularization was detected, respectively, between d1 and d4, and between d1 and d2. The vascular perfusion did not differ after d4 for endometrial tissue, whereas was reduced (P < .05) between d2 and d10 for mesometrium. After the first postpartum ovulation, an increase (P < .05) in vascular perfusion was observed from D0 to D5, followed by a decrease (P < .05) between D5 and D11 and an increase (P < .05) between D11 and D14. The novel vascular perfusion profile here described in the endometrium and mesometrium after ovulation is similar to the uterine vascular profile observed during estrous cycles and early pregnancy, indicating a fast return of the mare's uterus to cycling postpartum conditions.

Utility of indocyanine green (ICG) intra-operative angiography to determine uterine vascular perfusion at the time of radical trachelectomy

ObjectivesThe aim of our study was to measure and analyze uterine perfusion utilizing laser angiography with ICG during uterine artery sparing and non-sparing radical trachelectomy. MethodsData were collected from all patients diagnosed with early-stage cervical cancer that underwent laser angiography with ICG during open or laparoscopic radical trachelectomy from June 2012 to December 2015. Regression analysis was use to determine the p values and R-squares on fluorescence, surgical time, hospital stay, age and BMI; a p-value<0.05 was considered statistically significant. ResultsA total of 20 patients met the inclusion criteria and were included in this study. Ten patients underwent uterine artery-sparing surgery, and ten patients underwent uterine artery non-sparing surgery. The most frequent stage for the entire cohort was IA2 (55%), and the most common histologic subtype was squamous cell carcinoma (49%). Lymph-vascular invasion was noted in 30% of the patients. There was no statistical significance difference in the mean ICG fundal fluorescence intensity between the uterine artery-sparing group 162.5 (range, 137–188) and the uterine artery non-sparing group 160.5 (range, 135–186), p=0.22. In both groups, 100% of the patients regained their menstrual function by postoperative week 8. A total of 4 (40%) pregnancies have occurred in the uterine artery-sparing group and 3 (30%) in the non-uterine artery-sparing group. ConclusionsBased on our real-time intraoperative angiography observations, there is no need to preserve the uterine artery during radical trachelectomy to maintain uterine viability.

Correlación de la velocidad media de la arteria uterina con la edad gestacional como indicador de perfusión vascular uterina

Objetivo: Determinar la correlación de la velocidad media de la arteria uterina con la edad gestacional como indicador de perfusión vascular uterina. Diseño: Estudio transversal, analítico. Institución: Hospital III Honorio Delgado de Arequipa, Perú. Participantes: Gestantes entre 14 a 41 semanas de gestación, que acudieron al hospital para evaluación ecográfica de rutina sin evidencia de patología. Métodos: A gestantes entre 14 a 41 semanas de gestación que acudieron al hospital para evaluación ecográfica de rutina sin evidencia de patología, entre julio 2013 y abril 2014, se realizó ecografía Doppler de las arterias uterinas y se correlacionó la velocidad media (MnV) promedio de ambas arterias con la edad gestacional determinada por ecografía del primer trimestre. Se buscó la correlación de la MnV promedio en el segundo (14 a 28 semanas) y tercer (29 a 41 semanas) trimestres de gestación y se determinó el percentil 5 para dicha velocidad. Principales medidas de resultados: Cálculo de la MnV promedio de ambas arterias uterinas y correlación con la edad gestacional. Resultados: Se evaluó 138 gestantes con una edad materna promedio de 26,4 años y se encontró un coeficiente de correlación de la MnV promedio con la edad gestacional de r = 0,556 (r2 = 0,31). En el segundo y tercer trimestre de gestación el coeficiente de correlación fue r = 0,634 (r2 = 0,40) y r = 0,32 (r2 = 0,001), respectivamente. El percentil 5 de la MnV promedio en el tercer trimestre fue 67,50 cm/s. Conclusiones: Existió correlación de la MnV con la edad gestacional, cuyos valores menores al percentil 5 según la edad gestacional podrían ser utilizados como indicadores de hipoperfusión vascular uterina en gestantes del segundo y tercer trimestre.

Assessment of Uterine Vascular Perfusion During the Estrous Cycle of Mares in Connection to Circulating Leptin and Nitric Oxide Concentrations

The objective of this study was to find difference in vascular perfusion of uterine horns or uterine body throughout the estrous cycle and their relation to circulating nitric oxide and leptin concentrations. Five cyclic mares were subjected to transrectal Doppler ultrasonography and blood sampling for 18 days. Area of color and power Doppler modes was measured in pixels. Day (P = .0001) of the estrous cycle and ovulation (P = .0001) influenced uterine blood flow. Uterine body blood flow directed away from the transducer (blue, P = .0001) increased from day −5 until day 0 (day of ovulation), and its power (P = .0001) blood flow increased from day −6 until day 0; then, both decreased until days 12 and 10, respectively. Conversely to the contralateral uterine horn, ipsilateral uterine horn blood flow directed away from the transducer (blue, P = .0001) increased from day −5 until day −1, and its power (P = .0001) blood flow increased from day −6 until day 0; then, both decreased until day 10. Nitric oxide concentrations (P = .0001) attained two major peaks; the first on day −3 and the other persisted from day 2 until day 5. Leptin concentrations increased (P > .001) with a maximum value on day 0 and then decreased until a minimum value on day 9. In conclusion, during the estrous cycle, ipsilateral uterine horn and uterine body blood vessels had similar blood flow. Both leptin and nitric oxide played a role during follicle growth, ovulation, and corpus luteum development and modulated uterine blood flow before and after ovulation.

Real-time characterization of the uterine blood flow in mares before and after artificial insemination

The present experiment was divided into two studies to investigate the effect of age and endometrial degeneration on uterine blood flow of mares throughout the immediate post-breeding period. In study 1, uterine blood flow was characterized in mares (n=7 mares/group) with minimal, moderate or severe endometrial degenerative changes (GI, GII and GIII, respectively). In study 2, the effect of age was investigated using young (≤6 years) and old (≥15 years) mares (n=7 mares/group). Uterine vascular perfusion and mesometrial pulsatility index (PI) were evaluated every hour from H0 (moment immediately before AI) to H12. In study 1, a pronounced and transitory increase on uterine vascular perfusion was detected (P<0.001) between H1 and H3 in the three endometrial groups. In addition, GIII mares had greater (P<0.05) mesometrial PI than GI mares during the post-breeding period, denoting reduced uterine blood flow. In study 2, a transitory increase on uterine vascular perfusion was also observed in both age groups during the first hour after mating. However, mesometrial PI of young and old mares was similar (P>0.05) and constant (P>0.7) through the first 12h after AI. Results demonstrated, for the first time, the immediate changes on uterine vascular perfusion and mesometrial PI in response to semen infusion. Moreover, reduced blood flow of the uterus during the post-breeding period was strongly associated with endometrial degenerative changes in mares, regardless of age.

Effect of age and endometrial degenerative changes on uterine blood flow during early gestation in mares

The present experiment was divided into three studies to investigate the effect of age and endometrial degeneration on uterine blood flow of mares. In study 1, the influence of semen infusion and conception was evaluated using noninseminated (non-AI), inseminated nonpregnant (AI-NP), and inseminated pregnant (AI-P) mares (n = 7 mares/group). In study 2, the effect of age was investigated using young (≤6 years), adult (from 8 to 12 years), and old (≥15 years) pregnant mares (n = 7 mares/group). In study 3, uterine blood flow was also characterized in pregnant mares (n = 7 mares/group) with minimal, moderate, or severe endometrial degenerative changes (GI, GII, and GIII, respectively). Uterine vascular perfusion and pulsatility index from mesometrium attachment arteries were recorded daily from Day 0 (day of ovulation) to Day 12 and between Days 0 and 20 in nonpregnant and pregnant mares. Analysis according to the position of the embryo (uterine horn with embryo vs. opposite horn) was performed from Day 12 until Day 20. In study 1, increased uterine vascular perfusion and decreased pulsatility index were detected (P < 0.001) between Days 3 and 5 in non-AI and AI-P mares, whereas the uterine vascular perfusion of AI-NP mares did not change (P > 0.05) throughout the experiment. In study 2, the vascular perfusion of the uterine horn with embryo was higher (P < 0.001) than in the opposite uterine horn from Day 12 until Day 20 in both young and adult mares. With exception of Day 15, both uterine horns of old mares showed similar (P > 0.1) vascular perfusion from Day 12. In study 3, vascular perfusion of both uterine horns was lower (P < 0.001) in GIII mares than in GI and GII mares. After Day 15, the scores for uterine vascular perfusion were consistently greater (P < 0.001) in the uterine horn with embryo than in the opposite horn in GI mares. Results showed, for the first time, the reduced blood flow of the gravid uterus during early gestation in older mares and in mares with significant endometrial degeneration.

Effect of the conceptus on uterine vascular perfusion during early pregnancy of SCNT cloned bovine embryos.

Effect of the conceptus on uterine vascular perfusion during early pregnancy of SCNT cloned bovine embryos.

Influence of l-arginine supplementation on reproductive blood flow and embryo recovery rates in mares

Supplementation with l-arginine can increase uterine arterial blood flow and vascular perfusion of the preovulatory follicle in mares. Increased vascular perfusion of the preovulatory follicle has been correlated with successful pregnancy in mares. The objective of this study was to determine if supplemental l-arginine would increase ovarian arterial blood flow, vascular perfusion of the preovulatory follicle, and embryo recovery rates in mares. Mares were blocked by age and breed and assigned at random within block to l-arginine supplementation or control groups. Mares were fed l-arginine beginning 17 days before and through the duration of the study. Transrectal Doppler ultrasonography was used to measure ovarian arterial blood flow and vascular perfusion of the preovulatory follicle daily when it reached 35 mm and subsequent CL on Days 2, 4, and 6. Mares, on achieving a follicle of 35 mm or more were bred via artificial insemination and an embryo collection was attempted 7 days after ovulation. Treatment did not affect interovulatory interval (arginine-treated, 18.1 ± 2.6 days; control, 20.7 ± 2.3 days) or embryo recovery rate (arginine-treated, 54%; control, 48%). Mares treated with l-arginine had a larger follicle for the 10 days preceding ovulation than control mares (30.4 ± 1.2 and 26.3 ± 1.3 mm, respectively; P < 0.05) and vascular perfusion of the dominant follicle tended (P = 0.10) to be greater for the 4 days before ovulation. No differences were observed between groups in diameter or vascular perfusion of the CL. Resistance indices, normalized to ovulation, were not significantly different between groups during the follicular or luteal phase. Oral l-arginine supplementation increased the size and tended to increase perfusion of the follicle 1, but had no effect on luteal perfusion or embryo recovery rates in mares.

112 INFLUENCE OF LOW-VOLUME UTERINE FLUSHING ON UTERINE VASCULAR PERFUSION AND ENDOMETRIAL THICKNESS DURING EARLY DIOESTRUS IN BEEF CATTLE

The relevance of the uterine secretions during early embryonic development is due to its role as the only source of nutrients for embryo prior to implantation. Probing of the uterine secretions during early pregnancy without compromising gestation would be an important tool for the identification of fertility markers. Manipulations of the uterus could lead to a local increase in blood flow, which could result from the physical stimulation or tissue damage and inflammation. This study aimed to evaluate the effects of low-volume flushing of the uterine horn contralateral to the ovulation on uterine vascular perfusion and endometrial thickness 6 days after artificial insemination (AI). Transrectal B-mode and color Doppler ultrasonography (MyLab30 Vet Gold; Esaote Healthcare) were used to compare changes on endometrial thickness and uterine vascular perfusion, respectively. Examinations were carried out on 15 Nelore cows just prior to the flushing procedure (0 h), 6 and 24 h later. Cows were inseminated after oestrus detection and uterine flushings were performed 6 d after AI. The uterine horn, contralateral to the corpus luteum, was flushed with 20 mL of PBS using a Foley catheter. After massage of the uterine horn, flushing was collected in a syringe by suction. Vascular perfusion was estimated by scoring the extent of colored areas within the endometrium and mesometrium. Vascular perfusion scores indicated nil (1), minimal (2), intermediate (3), and maximal (4) vascular perfusion. Endometrial thickness was measured by taking the maximum diameter and its perpendicular diameter; these values were summed and then divided by four. Data that were not normally distributed were transformed to natural logarithms or ranks. For each variable, the main effect of side (flushed and nonflushed horns), hour, and their interaction were analyzed using the PROC MIXED procedure from the SAS software. The least significant difference method was used for the comparison of means among hours. An hour effect was detected for endometrial and mesometrial scores of vascular perfusion (Table 1), representing an increased vascular perfusion about 6 h after the flushing procedure and a return to basal perfusion at 24 h. Results indicate that the unilateral flushing procedure increases vascular perfusion on both uterine horns. For endometrial thickness, no effect of side, hour, or their interaction was detected. In conclusion, unilateral low-volume uterine flushing results in a transitory increase in uterine blood flow about 6 h post-flushing. Further studies are needed to evaluate other inflammatory characteristics and the potential effect on pregnancy rate in response to the uterine flushing procedure performed during early pregnancy. Table 1.Scores of endometrial and mesometrial vascular perfusion and endometrial thickness of both uterine horns at the time of uterine flushing (0 h), 6 and 24 h post-flushing (mean ± standard error of the mean)

Contrast-Enhanced Ultrasound Reveals Real-Time Temporospatial Changes in Uterine and Placental Vascular Perfusion During Early Pregnancy in Rhesus Macaques.

Classical morphological studies on preserved uterine tissue sections have provided an anatomical understanding of the uterine vascular changes that occur during early pregnancy, but little is known regarding the physiological changes (i.e. uterine blood flow) that occur in vivo during pregnancy initiation in primates. Doppler ultrasound (DUS), the prevailing technique used to detect the fetal heartbeat, lacks the sensitivity needed to measure the low velocity vascular perfusion associated with implantation and subsequent early placentation. Contrast-enhanced ultrasound (CEU) is a non-invasive method with high spatial resolution used to assess quantitative differences in vascular blood flow and volume. Thus, the goals of this study were to use CEU to characterize and quantify the temporal changes in the vascular adaptations associated with early pregnancy, and to compare the information provided by CEU and DUS. Adult female rhesus macaques (n = 5) in the time-mated breeding program at ONPRC were pair-caged with males of proven fertility for 96 hr during the window of ovulation, and mating was confirmed by video recording. DUS and CEU began 14 days after the first day of pairing (DAP) and continued every 3 days until a fetal heartbeat was detected by DUS. The uterus of sedated animals was first visualized on a Voluson Expert 730 system (GE HealthCare) with a 2D trans-abdominal probe with Doppler capability. CEU was performed with a contrast-specific imaging technique, via an Acuson Sequoia system (Siemens), during a continuous intravenous infusion of lipid microbubble contrast reagent (Definity). Vascular blood flow and blood volume were quantified for the myometrium, endometrium and placental lobes from replenishment kinetics on the intensity data obtained after a high power pulse sequence destroyed all microbubbles in the acoustical beam. A functional corpus luteum formed (P4 > 4 ng/mL) in all females and 80% (4/5) became pregnant, as confirmed by a fetal heartbeat observed on 28.0±1.2 DAP. CEU clearly delineated the primary and secondary lobes of the placenta ~2 days apart (paired t-test; P<0.05), on 19.8±1.0 and 22.0±1.2 DAP, respectively. DUS was not able to differentiate between the placental lobes and detected endometrial thickening on 22.8±1.0 DAP, 3 days after (P<0.05) CEU identified the primary placental lobe. Visualization of the primary lobe correlated with the vaginal bleeding associated with implantation. The CEU calculated vascular perfusion was greater (P<0.05) in the primary placental lobe, which exhibited a 29% increase in relative blood volume and a 40% increase in relative blood flow, compared with the secondary lobe on the day of heartbeat detection. CEU identified greater flow through the hypertrophied endometrial spiral arteries supplying the placental lobes compared with other regions of the endometrium, whereas changes in the myometrium were not seen. In sum, CEU has defined the temporospatial changes in vascular perfusion associated with the anatomical vascular remodeling during early pregnancy in primates. Thus, CEU has great potential as a tool for basic researchers examining the peri-implantation period in the primate endometrium. Furthermore, CEU has promise as a future application for obstetricians, to resolve vascular complications that arise during pregnancy, including ectopic implantation and preeclampsia. Supported by NIH grants HD055744, HD18185, HD07133 and RR000163. (platform)

Uterine vascular perfusion and spectral-Doppler measurements during early gestation in mares: new concepts of evaluation

Uterine vascular perfusion and spectral-Doppler measurements during early gestation in mares: new concepts of evaluation

Local effect of the conceptus on uterine vascular perfusion during early pregnancy in heifers

Colour-Doppler ultrasonography was used to study the spatial relationship between vascular perfusion in the middle of each uterine horn and the reported location of the embryo proper and expanding conceptus using endometrial vascularity scores 1-4 (nil-maximal). Vascularity increased in both uterine horns between days 14 and 18 (day 0=ovulation) in nonpregnant heifers (n=6) but not in pregnant heifers (n=11). The increase was temporally associated with decreasing plasma progesterone and increasing oestradiol. In pregnant heifers, a transient increase in endometrial vascularity in the ipsilateral horn (horn with embryo) was not detected before day 18, despite a reported transient increase in blood flow in the ipsilateral uterine artery between days 13 and 17. Endometrial vascularity in the ipsilateral horn first increased (P<0.05) between days 18 and 20. Day 20 is the reported day of adhesiveness between chorion and uterus. An increase (P<0.05) in the contralateral horn between days 18 and 22 was slight, but a greater increase occurred after day 32. Day 32 is the reported day of entry of the allantochorion into the contralateral horn. By day 42, scores were similar between the two horns, and the allantochorion reportedly fills both horns. On days 42-60, at a time when placentomes apparently are limited to the ipsilateral horn, vascularity remained elevated in the ipsilateral horn but decreased in the contralateral horn. Results support the hypothesis that vascular perfusion in each uterine horn during early pregnancy is mediated by direct contact between conceptus and uterus.