Abstract
Background:
Cardioplegia and cardiopulmonary bypass (CP/CPB) alters coronary arteriolar response to thromboxane A2 (TXA2) in patients undergoing cardiac surgery. Comorbidities, including hypertension (HTN), can further alter coronary vasomotor tone. This study investigates the effects of HTN on coronary arteriolar response to TXA2 pre- and post-CP/CPB and cardiac surgery.
Materials and Methods:
Coronary arterioles pre- and post-CP/CPB were dissected from atrial tissue samples in patients with no HTN (NH, n=9), well-controlled HTN (WC, n=12), or uncontrolled HTN (UC, n=12). In-vitro coronary microvascular reactivity was examined in the presence of TXA2 analog U46619 (10−9–10−4M). Protein expression of TXA2 receptor in the harvested right atrial tissue samples were measured by immunoblotting.
Results:
TXA2 analog U46619 induced dose-dependent contractile responses of coronary arterioles in all groups. Pre-CPB contractile responses to U46619 were significantly increased in microvessels in the UC group compared to the NH group (p<0.05). The pre-CP/CPB contractile responses of coronary arterioles were significantly diminished post-CP/CPB among the three groups (p<0.05), but there remained an increased contractile response in the microvessels of the UC group compared to the WC and NH groups (p<0.05). There were no significant differences in U46619-induced vasomotor tone between patients in the NH and WC groups (p>0.05). There were no differences in expression of TXA2R among groups.
Conclusions:
Poorly controlled HTN is associated with increased contractile response of coronary arterioles to TXA2. This alteration may contribute to worsened recovery of coronary microvascular function in patients with poorly controlled hypertension after CP/CPB and cardiac surgery.
Keywords: hypertension, coronary, microvascular, thromboxane A2, cardiopulmonary bypass
INTRODUCTION:
Cardiac surgery with cardioplegia and cardiopulmonary bypass (CP/CPB) alters coronary microvascular tone, which directly affects myocardial perfusion in the peri-operative setting.1–3 Furthermore, patients undergoing cardiac surgery often have several co-morbidities, such as diabetes and hypertension, that further dysregulate coronary microvascular function.4 The combination of comorbidity-induced microvascular dysfunction with the physiologic alterations that occur with CP/CPB may result in compounded dysregulation and malperfusion.4–8
Hypertension is one important co-morbidity in patients undergoing cardiac surgery that impacts coronary microvascular function.4,9,10 Despite the availability of multiple antihypertensive agents, and recent guidelines to titrate these agents in patients with cardiovascular disease for a goal blood pressure of less than 130/80, a substantial proportion of patients remain uncontrolled, leading to persistent cardiovascular risk.11 We have previously shown that uncontrolled hypertension is associated with dysregulated coronary myogenic tone and vasoconstrictive response to phenylephrine before and after CP/CPB.8 However, studies are lacking on the effects of hypertension control on other physiologic pathways involved in coronary microvascular function.
Thromboxane A2 (TXA2), a potent endothelial-derived vasoconstrictor and platelet activator, may contribute to coronary microvascular dysregulation in the setting of CP/CPB. TXA2 is derived from arachidonic acid and activates vascular smooth muscle contraction via activation of phospholipase C, and its activity can be further regulated by phospholipase A.12 We have previously demonstrated decreased contractile response of coronary arterioles to a thromboxane A2 (TXA2) analog following CP/CPB, which was independent of TXA2 receptor expression.13 TXA2 is known to play a role in the pathophysiology of hypertension and other cardiovascular disease,14 though the relationship between hypertension and TXA2-mediated coronary microvascular function is poorly investigated. Understanding this relationship is particularly important in the context of cardiac surgery with CP/CPB given the role of coronary microvascular function in myocardial perfusion in the peri-operative setting.
Therefore, the goal of this study is to investigate differences in coronary microvascular vasomotor responses to thromboxane A2 before and after CP/CPB in patients with no hypertension, well-controlled hypertension, and uncontrolled hypertension. We hypothesized that the contractile response to TXA2 would be increased in patients with poorly controlled hypertension, possibly via altered TXA2 signaling pathways.
Material and Methods:
All procedures were approved by the Institutional Review Board (IRB) of Rhode Island Hospital, Alpert Medical School of Brown University (IRB #004410, 03/10/10), and informed consent was obtained from all enrolled patients prior to tissue collection and involvement in the study as required by the IRB.
Patient Groups
Patient groups were divided based on hypertension (HTN) status. HTN status was determined by documented history of HTN, use of prescription anti-hypertensive agents, and average systolic blood pressure (SBP) measurements over one year prior to cardiac surgery. Patients with no documented active HTN, no use of prescription anti-hypertensive agents, and an average SBP less than 130 mmHg were placed in the non-hypertensive (NH) group. Patients with a documented history of HTN, use of prescription anti-hypertensive agents, and an average SBP less than 130 mmHg were placed in the well-controlled HTN (WC) group. Patients with a documented history of HTN, with or without use of prescription anti-hypertensive agents, and an average SBP greater than 130 mmHg were placed in the uncontrolled HTN (UC) group. Prescription anti-hypertensive agent use was documented as of the patient’s most recent history and physical documentation. At our institution, patients are instructed to hold angiotensin-converting enzyme inhibitors and angiotensin receptor blockers 48 hours prior to surgery, and to continue beta blockers and calcium channel blockers through the morning of surgery. Patients without coronary artery disease are instructed to hold aspirin for five days prior to surgery, while those with coronary artery disease are instructed to continue home aspirin until the time of surgery, and these patients receive post-operative aspirin starting the day of the procedure.
Tissue Collection
Right atrial appendage tissue was collected from patients undergoing cardiac surgery with CP/CPB before and after CPB. Cold blood CP solution (4° to 8°C) consisted of a 4:1 mixture of oxygenated blood and hyperkalemic crystalloid CP solution (CAPS, Lanham, MD). An initial 600 to 1,000 ml (introduction) of cold-blood (4° to 8°C) hyperkalemic (K+ 25 mmol/L) CP solution was delivered antegrade into the aortic root, followed by 200 to 500 mL (8 mmol/L, maintenance) of cold blood CP solution every 15 to 20 minutes.
Two samples were obtained using a double-purse-string technique with 3–0 polypropylene sutures. The first sample was collected before CPB initiation during venous cannula placement into the right atrium. During collection, the superior suture was tightened to secure the cannula, while the inferior suture remained loose to expose the atrial tissue to cardioplegia solution, CPB, and reperfusion. The second sample was collected between the two purse-string sutures after removal of cross clamp removal and termination of CPB, but prior to administration of protamine. Samples designated for immunoblotting were frozen in liquid nitrogen, and samples designated for in-vitro microvascular reactivity studies were stored in cold Krebs buffer.
In-vitro Coronary Microvascular Studies
Coronary arterioles (80–180μm internal diameter) were harvested from atrial tissue from NH (n=5), WC (n=8), and UC (n=8) groups, and dissected using a dissecting microscope. Microvessels were placed in a microvessel chamber containing circulating warm (37 degrees Celsius) and oxygenated (95% oxygen and 5% carbon dioxide) Krebs buffer solution, cannulated with dual glass micropipettes (30–80μm in diameter), and secured in place with 10–0 nylon monofilament sutures. Microvessels were then pressurized to 40 mmHg in a no-flow state using a burette manometer filled with Krebs buffer solution. The microvessel image was projected onto a monitor using an inverted microscope (40–200x, Olympus CK2, Olympus Optical) connected to a video camera. Internal luminal diameter was measured with an electronic dimension analyzer. Microvessels were bathed in Krebs buffer in the microvessel chamber for at least 30 minutes, followed by application of TXA2 analog U46619 (10−9–10−4M) and assessment of in-vitro contractile responses by vessel myography.
Immunoblotting
Atrial tissue samples from NH (n=4), WC (n=4), and UC (n=4) groups were dissected and cleaned of connective tissues and solubilized in RIPA buffer. Protein concentration was determined using a Pierce BCA Protein Assay Kit (ThermoFisher Scientific, Waltham, MA), and 35μg of total protein was fractionated onto a 4–12% Bis-Tris gel (ThermoFisher Scientific), followed by transfer to a nitrocellulose membrane (ThermoFisher Scientific). Membranes were then incubated at 4 degrees Celcius overnight with a 1:500 dilution of rabbit polyclonal primary antibody to TXA2R, 1:1000 dilution of mouse polyclonal primary antibody to phospholipase A2 receptor (PLA2R), and 1:1000 dilutions of rabbit polyclonal primary antibody to phospholipase beta 3 (PLCß3) and phospholipase gamma 1 (PLCγ1) (Abcam, Cambridge, UK). Membranes were then rinsed and incubated with goat polyclonal anti-rabbit or anti-mouse secondary antibodies for one hour at room temperature. Membranes were then rinsed, incubated in chemiluminescent detection solution, and captured with a digital camera system (Bio-Rad ChemiDoc MP, Life Science, Hercules, CA). All membranes were probed with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Cell Signaling, Danvers, MA) to correct for loading error. NIH Image J software was used to perform densitometric analysis of band intensity. Complete blot images are shown in the Supplemental Figure.
Immunofluorescence
Immunofluorescence studies were performed as previously described.7 Briefly, paraffin-embedded atrial tissue sections were de-paraffinized, antigen unmasked using sodium citrate solution, blocked using 3% BSA, and incubated with primary antibodies to α-smooth muscle actin (α-SMA) and TXA2R (Abcam, Cambridge, UK) or no additional antibody for negative control. Sections were then incubated with Alexa-fluor tagged secondary antibodies (Abcam, Cambridge, UK). Images were captured at 20X magnification with an Olympus VS200 Slide Scanner.
Data Analysis
Microvessel responses to TXA2 analog U46619 are expressed as percent constriction compared to baseline. Microvascular reactivity data were analyzed using 2-way repeated measures ANOVA followed by Tukey’s multiple comparison’s test when appropriate. Western blot data was tested for normality using Shapiro-Wilk testing, then analyzed using paired t tests if normal or Wilcoxon signed-rank test if non-normal, with Bonferroni corrections to compare densitometry of samples before and after CPB. Data was analyzed using unpaired t tests if normal or Wilcoxon rank sum test if non-normal with Bonferroni corrections to compare densitometry of samples from NH vs WC vs UC patients pre-bypass or post-bypass. Correlation data was analyzed using Spearman’s test. Data are reported as mean and standard deviation (SD). Probability values <0.05 were considered statistically significant.
RESULTS:
Patient Characteristics
Tissue samples from a total of 33 patients were studied, with 9 patients in the NH group (5 for microvessel studies, 4 for immunoblotting), 12 patients in the WC group (8 for microvessel studies, 4 for immunoblotting), and 12 patients in the UC group (8 for microvessel studies, 4 for immunoblotting). Patients underwent a variety of cardiac surgical procedures, the majority of which included CABG. Mean age was 63 ± 10 years old in the NH group, 65 ± 10 years old in the WC group, and 68 ± 7 years old in the UC group. Gender distribution was 8 males and 1 female in the NH group, 11 males and 1 female in the WC group, and 10 males and 2 females in the UC group. Patients from each group underwent a variety of cardiac surgical procedures with the majority being coronary artery bypass grafting. The average clinic systolic BP in mmHg was 119 ± 10 in the NH group, 117 ± 9 in the WC group, and 137 ± 5 in the UC group. Complete patient characteristics, including classes of anti-hypertensive drugs prescribed to patients as of their most recent pre-operative history and physical documentation are listed in Table 1. One patient in the NH group was prescribed a beta blocker for an indication other than hypertension. Of note, all patients who were not previously on beta blockers were started on beta blockers pre-operatively for cardioprotection. Among the patients from which samples were collected for microvessel analysis, patients in the UC group tended to have higher systolic blood pressures in the immediate pre-operative time period compared to the other groups (NH 123.2±14.8, WC 141.9±25.2, UC 162.3±17.9). There were no significant differences in duration of post-operative inotrope use among groups (NH 1732±2865 min, WC 1535±1146 min, UC 4074± 6038.5, p=0.25).
Table 1:
Baseline characteristics.
| Baseline Characteristics | No HTN (n=9) | Controlled HTN (n=12) | Uncontrolled HTN (n=12) |
|---|---|---|---|
| Age - yr. | 63 ± 10 | 65 ± 10 | 68 ± 7 |
| Male - n. (%) | 8 (67%) | 11 (92%) | 10 (83%) |
| BMI (kg/m2) | 27.6 ± 5.7 | 32.4 ± 7.0 | 31.6 ± 5.1 |
| HbA1c (%) | 6.2 ± 1.6 | 6.9 ± 1.3 | 6.3 ± 0.8 |
| CABG only - n. (%) | 7 (78%) | 6 (50%) | 8 (67%) |
| Valve surgery only - n. (%) | 1 (11%) | 3 (25%) | 1 (8%) |
| CABG + valve surgery - n. (%) | 1 (11%) | 3 (25%) | 3 (25%) |
| CPB Time (min) | 92 ± 47 | 141 ± 57 | 100 ± 61 |
| Cross Clamp Time (min) | 71 ± 40 | 107 ± 43 | 81 ± 51 |
| Systolic BP (mmHg) | 119 ± 10 | 117 ± 9 | 137 ± 5 |
| Diastolic BP (mmHg) Anti-hypertensive drug class | 72 ± 5 | 69 ± 6 | 77 ± 7 |
| ACEi - n (%) | 0 (0%) | 5 (42%) | 5 (42%) |
| ARB - n (%) | 0 (0%) | 2 (17%) | 5 (42%) |
| CCB - n (%) | 0 (0%) | 3 (25%) | 3 (25%) |
| Beta Blocker - n (%) | 1 (11%) | 10 (83%) | 8 (67%) |
| Thiazide - n (%) | 0 (0%) | 3 (25%) | 3 (25%) |
| Loop Diuretic - n (%) | 0 (0%) | 3 (25%) | 0 |
Baseline characteristics listed for patients undergoing cardiac surgery with cardioplegia/cardiopulmonary bypass from whom right atrial appendage tissue was collected. Hypertension (HTN) status determined by documented history of HTN, use of prescription anti-hypertensive agents, and average systolic blood pressure measurements over one year prior to cardiac surgery. Continuous variables presented as mean ± standard deviation; Abbreviations: BMI = body mass index; HbA1c = hemoglobin A1c; CABG = coronary artery bypass surgery; CPB = cardiopulmonary bypass; BP = blood pressure; ACEi = angiotensin converting enzyme inhibitor; ARB = angiotensin II receptor blocker; CCB = calcium channel blocker.
Microvascular Reactivity
TXA2 analog U46619 induced dose-dependent contractile responses of coronary arterioles in all three groups. Before CP/CPB, contractile responses to U46619 were significantly increased in microvessels in the UC group compared to those of the NH group (p<0.05) with trends towards increased contractile response compared to the WC group (p<0.10 at concentrations greater than 10−6M). The pre-CP/CPB contractile responses of coronary arterioles were significantly diminished post-CP/CPB among the three groups (p<0.05), but there remained an increased contractile response in the microvessels of the UC group compared to those in the WC and NH groups (p<0.05). There were no significant differences in U46619-induced vasomotor tone between patients in the NH and WC groups (p>0.05) (Figure 1). There was no correlation between U46619-induced vasomotor tone post-CPB and post-operative inotrope use duration (Rs=0.22, p=0.35).
Figure 1: In-vitro constriction response of coronary arterioles to thromboxane A2 analog before and after cardiopulmonary bypass.
Coronary arteriolar constriction response to thromboxane analog U46619 (10−9–10−4 M) in patients with no hypertension (HTN) (NH, n=5), well-controlled HTN (WC, n=8), and poorly controlled HTN (UC, n=8) (A) pre- and (B) post-cardiac surgery with cardioplegia/cardiopulmonary bypass (CP/CPB). Constriction responses within each HTN group pre- and post- CP/CPB are also shown (C-E). Arterioles were harvested from right atrial appendage tissue. HTN status determined by documented history of HTN, use of prescription anti-hypertensive agents, and average systolic blood pressure measurements over one year prior to cardiac surgery. Data plotted as mean ± SD. *p<0.05 pre-CP/CPB vs post-CP/CPB. #p<0.05 UC vs NH. +p<0.05 UC vs NH and UC vs WC.
Protein Expression
There were no significant differences in expression of TXA2R from pre- to post-CPB in the NH (p=0.31), WC (p>0.5), and UC (p=0.5) groups. There were no significant differences in expression of TXA2R at baseline or after CP/CPB between groups. Pre- to post-CPB expression of PLA2R and PLCß3 was unchanged among all three groups, while expression of PLCγ1 was decreased from pre- to post-CPB in the WC group (p=0.036). There were no significant differences in baseline or post-CPB expression of PLA2R and PLCγ1 among the groups. There was decreased expression of PLCß3 in post-CPB samples from the WC group compared to the NH group (p=0.032) (Figure 2).
Figure 2: Protein expression related to thromboxane A2 signaling.
Immunoblot results show expression of thromboxane A2 receptor (TXA2R), phospholipase Cß3 (PLCß3), phospholipase Cγ1 (PLCγ1), and phospholipase A2 receptor (PLA2R) in right atrial tissue samples from patients with no hypertension (HTN) (NH, n=4), well-controlled HTN (WC, n=4), and poorly controlled HTN (UC, n=4), before (pre-) and after (post-) cardiac surgery with cardioplegia/cardiopulmonary bypass (CP/CPB). HTN status determined by documented history of HTN, use of prescription anti-hypertensive agents, and average systolic blood pressure measurements over one year prior to cardiac surgery. Data presented as fold changes in optical density compared to average optical density in NH group. Complete blots provided in Supplemental Figure. Upper and lower borders of box represent upper and lower quartiles, middle horizontal line represents median, upper and lower whiskers represent maximum and minimum values of non-outliers. *p<0.05 pre-CPB vs post-CPB. #p<0.05 NH post-CPB vs WC post-CPB.
Protein Localization
Immunofluorescent studies confirmed that TXA2R was expressed on coronary microvessels as stained by α-SMA (Figure 3).
Figure 3: Localization of TXA2R in coronary microvessels.
Immunofluorescent staining of human atrial tissue sections in patients undergoing cardiac surgery with cardioplegia / cardiopulmonary bypass demonstrated expression of thromboxane A2 receptor (TXA2R, red) at coronary microvessels as stained by α-smooth muscle actin (α-SMA, green). Nuclei stained with DAPI (blue). Negative control stained only with α-SMA showed low level of background fluorescence without TXA2R signal. Images obtained at 20X magnification.
DISCUSSION:
In the present study, we found that in coronary arterioles isolated from human atrial tissue samples, the constrictive response to TXA2 is enhanced in patients with poorly controlled hypertension compared to patients with no HTN and well-controlled HTN. These effects were demonstrated prior to CP/CPB, and persistent after CP/CPB despite blunting of the vasoconstrictive response to TXA2 across patient groups. These findings were not associated with changes in expression of TXA2 receptor or phospholipases involved in TXA2-related signal transduction, which may be indicative of a change in functionality of the TXA2 receptor or its downstream mediators rather than changes in protein abundance.
The coronary microvasculature is made up of vessels <200 μm including capillaries and arterioles.4 One of the critical functions of coronary microvessels is regulation of vasomotor tone which directly affects coronary blood flow and myocardial perfusion.4 Normal regulation of coronary microvascular tone involves a complex interplay between endothelial-dependent and independent vasodilators and vasoconstrictors.4 One potent endothelial-derived vasoconstrictor is TXA2, which is derived from arachidonic acid following PLA2-catalyzed release from membrane phospholipids.12 TXA2 is released by platelets and endothelial cells and acts on TXA2 receptors on vascular SMC, activating a number of downstream mediators including PLC isoforms to increase intracellular calcium and promote vessel contraction.12
Cardioplegia with cardiopulmonary bypass is known to disrupt the normal vasomotor response to a number of vasomotor regulators, including TXA2.2,7,13,15 Coronary microvessels treated with a TXA2 analog have been shown to have reduced contractile responses after CP/CPB compared to baseline.13 Interestingly, in the current study, despite longer ischemic times in the WC group, post-CP/CPB arteriolar reactivity to TXA2 was decreased to a similar extent in the NH and UC group. It is plausible that arteriolar reactivity to TXA2 may decrease further with prolonged time on CPB, but this may not necessarily be a linear relationship and perhaps the greatest effect on decreased arteriolar reactivity is in the initial time period of CPB with tapering effects over time.
Furthermore TXA2 has been implicated in increased vasoconstriction and blood pressure in hypertension in small animal studies.14 There is evidence to suggest that TXA2 contributes to the development and maintenance of hypertension, and that hypertension itself can dysregulate TXA2 physiology.12,16–18 However, to our knowledge, the specific functional alterations in coronary microvasculature have not been previously investigated. This study demonstrated that in the presence of a TXA2 analog, coronary microvessels have an enhanced vasoconstrictive response in the setting of poorly controlled hypertension which is persistent despite CP/CPB, suggesting that poor blood pressure control may alter coronary arteriolar response to TXA2. These findings are clinically relevant, as increased vasoconstriction at the coronary microvasculature may reduce myocardial perfusion and contribute to vasospasm. These are novel findings on the effects of hypertension control on the coronary microvascular vasomotor tone, an area which has been to date poorly investigated. Importantly, there are multiple physiologic mediators of coronary microvascular tone, and poorly controlled hypertension likely dysregulates multiple pathways. Our group has recently demonstrated that poorly controlled hypertension increases coronary myogenic tone and vasoconstrictive response to phenylephrine.8 We assessed whether post-CPB vasoconstrictive response to TXA2 correlated with post-operative inotrope use duration, though there were no differences, nor were there significant differences in post-operative inotrope use duration among the three groups. However, limited sample size and patient variability may have confounded this analysis, and further studies are warranted to determine the clinical implications of pre-operative HTN control in cardiac surgery.
One important consideration is whether the effects of hypertension control on coronary arteriolar reactivity to TXA2 in this study is related to long-term control or shorter-term pre-operative blood pressure. We found that patients in the UC group tended to have higher systolic blood pressures compared to the other groups. It is unsurprising that the WC group tended to have higher SBP’s than the NH group given that ACEi and ARB’s are held in these patients 48 hours pre-operatively. The increased pre-operative blood pressure in the UC vs NH group is consistent with clinic blood pressure differences, thus prohibiting determination of whether our microvessel findings were secondary to an acute vs chronic effect. On the other hand, the WC group did tend to have higher pre-operative systolic blood pressures compared to the NH group, while the clinic blood pressures were comparable across these two groups, however microvessel reactivity to TXA2 was statistically similar between these two groups, which could be more suggestive of a chronic effect rather than acute. Therefore, it is likely that the findings in our study are related to long-term effects of hypertension control on coronary arteriolar reactivity to TXA2, though further studies may be warranted to investigate this in further detail.
We also sought to investigate specific molecular mechanisms by which hypertension status altered coronary arteriolar responsiveness to TXA2. One potential mediator we investigated was thromboxane A2 receptor, which is located in vascular smooth muscle cells to induce vasoconstriction.19 We found no differences in TXA2 receptor expression in patients with poorly controlled hypertension compared to other groups that would explain the differences in in-vitro microvessel reactivity to TXA2. Further, there were no differences in expression of proteins involved in TXA2 signaling, including PLA2R, PLCß3, and PLCG1 in the uncontrolled HTN group compared to the other groups. Therefore, differences in reactivity to TXA2 are less likely secondary to changes in expression of TXA2 receptor and its mediators, and perhaps due to changes in functionality of the receptor and/or its downstream mediators.
There are several limitations in this study to consider. One limitation is low sample size, particularly in our western blot analysis, in order to compare protein expression across six different groups on a single gel. Therefore the immunoblot studies may be underpowered to detect significant differences in relevant protein expression, especially given the necessity to statistically adjust for multiple comparisons. Another potential limitation is that most patients in all groups were on pre-operative aspirin, which is a known inhibitor of TXA2 production. Other studies suggest that aspirin administration increases the constrictive response to TXA2 analogs in-vitro.20 However, given that pre-operative aspirin use was similar across groups, and in fact slightly higher in the normotensive group compared to the uncontrolled hypertension group, we would not anticipate that aspirin use significantly altered the comparative analysis of TXA2 response across groups in this study. Finally, there are limitations of the study secondary to the clinical heterogeneity involved in analyzing human tissue samples. We collected and organized basic patient characteristics including age, gender, BMI, and HbA1c, however there may be other co-morbidities or clinical characteristics contributing to coronary microvascular tone and protein expression which are unmeasured.
CONCLUSIONS:
Our findings indicate that poorly controlled hypertension is associated with increased contractile response of coronary arterioles to TXA2, both pre- and post-cardioplegia and cardiopulmonary bypass in the setting of cardiac surgery. These alterations may have important implications on myocardial perfusion and recovery in the peri-operative setting and warrant further investigation.
Supplementary Material
Supplementary Figure: Complete Western blot images. Complete Western blot images for expression of thromboxane A2 receptor (TXA2R), phospholipase Cß3 (PLCß3), phospholipase Cγ1 (PLCγ1), and phospholipase A2 receptor (PLA2R) in right atrial tissue of patients with no hypertension (HTN) (NH, n=4), well-controlled HTN (WC, n=4), and poorly controlled HTN (UC, n=4), before (pre-) and after (post-) cardiac surgery with cardioplegia/cardiopulmonary bypass (CP/CPB). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control.
ACKNOWLEDGEMENTS:
We would like to thank all the nurses, physician assistants and perfusionists working in cardiac surgery at Rhode Island Hospital for collecting the tissue samples and recording patient characteristics. We would also like to thank the nurses and physician assistants in the Division of Cardiac Surgery of Rhode Island Hospital for collecting patient consent forms.
Funding:
This study was supported by the National Heart, Lung, and Blood Institute (NHLBI) 1F32HL160063-01 (S.A.S.); R01HL46716 and R01HL128831-01A1 (F.W.S.), 1R01HL127072-01A1, 1R01 HL136347-01, and R01HL136347-04S1 (J.F).
Footnotes
Disclosures: None
Presentation: Presented at the 18th Annual Academic Surgical Congress, Feb. 7–9, Houston, TX.
Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain
REFERENCES
- 1.Ruel M, Khan TA, Voisine P, Bianchi C, Sellke FW. Vasomotor dysfunction after cardiac surgery. Eur J Cardio-Thorac Surg Off J Eur Assoc Cardio-Thorac Surg. 2004. Nov;26(5):1002–14. [DOI] [PubMed] [Google Scholar]
- 2.Sellke FW, Friedman M, Dai HB, Shafique T, Schoen FJ, Weintraub RM, et al. Mechanisms causing coronary microvascular dysfunction following crystalloid cardioplegia and reperfusion. Cardiovasc Res. 1993. Nov;27(11):1925–32. [DOI] [PubMed] [Google Scholar]
- 3.Sodha NR, Feng J, Clements RT, Bianchi C, Boodhwani M, Ramlawi B, et al. Protein kinase C alpha modulates microvascular reactivity in the human coronary and skeletal microcirculation. Surgery. 2007. Aug;142(2):243–52. [DOI] [PubMed] [Google Scholar]
- 4.Sabe SA, Feng J, Sellke FW, Abid MR. Mechanisms and Clinical Implications of Endothelium-dependent Vasomotor Dysfunction in Coronary Microvasculature. Am J Physiol Heart Circ Physiol. 2022. Mar 25; [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Feng J, Chu LM, Dobrilovic N, Liu Y, Singh AK, Sellke FW. Decreased coronary microvascular reactivity after cardioplegic arrest in patients with uncontrolled diabetes mellitus. Surgery. 2012. Aug;152(2):262–9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Feng J, Anderson K, Singh AK, Ehsan A, Mitchell H, Liu Y, et al. Diabetes Upregulation of Cyclooxygenase 2 Contributes to Altered Coronary Reactivity After Cardiac Surgery. Ann Thorac Surg. 2017. Aug;104(2):568–76. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Sellke N, Gordon C, Lawandy I, Gorvitovskaia AY, Scrimgeour LA, Fingleton JG, et al. Impaired coronary contraction to phenylephrine after cardioplegic arrest in diabetic patients. J Surg Res. 2018. Oct;230:80–6. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Sabe SA, Kononov MA, Bellam KG, Sodha N, Ehsan A, Jackson WF, et al. Poorly controlled hypertension is associated with increased coronary myogenic tone in patients undergoing cardiac surgery with cardiopulmonary bypass. J Thorac Cardiovasc Surg. 2022. Aug 2;S0022–5223(22)00813–3. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Shah RM, Zhang Q, Chatterjee S, Cheema F, Loor G, Lemaire SA, et al. Incidence, Cost, and Risk Factors for Readmission After Coronary Artery Bypass Grafting. Ann Thorac Surg. 2019. Jun;107(6):1782–9. [DOI] [PubMed] [Google Scholar]
- 10.Terada T, Johnson JA, Norris C, Padwal R, Qiu W, Sharma AM, et al. Severe Obesity Is Associated With Increased Risk of Early Complications and Extended Length of Stay Following Coronary Artery Bypass Grafting Surgery. J Am Heart Assoc Cardiovasc Cerebrovasc Dis. 2016. Jun 1;5(6):e003282. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Whelton PK, Carey RM, Aronow WS, Casey DE, Collins KJ, Dennison Himmelfarb C, et al. 2017 ACC/AHA/AAPA/ABC/ACPM/AGS/APhA/ASH/ASPC/NMA/PCNA Guideline for the Prevention, Detection, Evaluation, and Management of High Blood Pressure in Adults: Executive Summary: A Report of the American College of Cardiology/American Heart Association Task Force on Clinical Practice Guidelines. Hypertens Dallas Tex 1979. 2018. Jun;71(6):1269–324. [DOI] [PubMed] [Google Scholar]
- 12.Nakahata N Thromboxane A2: physiology/pathophysiology, cellular signal transduction and pharmacology. Pharmacol Ther. 2008. Apr;118(1):18–35. [DOI] [PubMed] [Google Scholar]
- 13.Feng J, Liu Y, Chu LM, Clements RT, Khabbaz KR, Robich MP, et al. Thromboxane-induced contractile response of human coronary arterioles is diminished after cardioplegic arrest. Ann Thorac Surg. 2011. Sep;92(3):829–36. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Sellers MM, Stallone JN. Sympathy for the devil: the role of thromboxane in the regulation of vascular tone and blood pressure. Am J Physiol Heart Circ Physiol. 2008. May;294(5):H1978–1986. [DOI] [PubMed] [Google Scholar]
- 15.Robich MP, Araujo EG, Feng J, Osipov RM, Clements RT, Bianchi C, et al. Altered coronary microvascular serotonin receptor expression after coronary artery bypass grafting with cardiopulmonary bypass. J Thorac Cardiovasc Surg. 2010. Apr;139(4):1033–40. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Hu J, Yang Z, Chen X, Kuang S, Lian Z, Ke G, et al. Thromboxane A2 is involved in the development of hypertension in chronic kidney disease rats. Eur J Pharmacol. 2021. Oct 15;909:174435. [DOI] [PubMed] [Google Scholar]
- 17.Stier CT, Itskovitz HD. Thromboxane A2 and the development of hypertension in spontaneously hypertensive rats. Eur J Pharmacol. 1988. Jan 27;146(1):129–35. [DOI] [PubMed] [Google Scholar]
- 18.Taube C, Höhler H, Lorenz S, Förster W. The role of TXA2 in hypertension. Biomed Biochim Acta. 1984;43(8–9):S208–211. [PubMed] [Google Scholar]
- 19.Dorn GW, Becker MW. Thromboxane A2 stimulated signal transduction in vascular smooth muscle. J Pharmacol Exp Ther. 1993. Apr;265(1):447–56. [PubMed] [Google Scholar]
- 20.Toda N Mechanism of action of carbocyclic thromboxane A2 and its interaction with prostaglandin I2 and verapamil in isolated arteries. Circ Res. 1982. Dec;51(6):675–82. [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Supplementary Materials
Supplementary Figure: Complete Western blot images. Complete Western blot images for expression of thromboxane A2 receptor (TXA2R), phospholipase Cß3 (PLCß3), phospholipase Cγ1 (PLCγ1), and phospholipase A2 receptor (PLA2R) in right atrial tissue of patients with no hypertension (HTN) (NH, n=4), well-controlled HTN (WC, n=4), and poorly controlled HTN (UC, n=4), before (pre-) and after (post-) cardiac surgery with cardioplegia/cardiopulmonary bypass (CP/CPB). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control.