Lowered pulmonary arterial pressure prevents edema after endotoxin in sheep

Escherichia coli endotoxin causes increased capillary membrane permeability and increased pulmonary arterial pressure (PAP) in sheep. If the pulmonary hypertension extends to the level of the microvasculature, then the increased microvascular pressure may contribute to the pulmonary edema caused by endotoxin. We tested the hypothesis that reducing the pulmonary hypertension would reduce the amount of edema caused by endotoxin. Twelve sheep were chronically instrumented with catheters to measure PAP, left atrial pressure, and central venous pressure. The sheep were divided into two groups. One group (E) of six sheep received an intravenous infusion of 4 micrograms/kg of E. coli endotoxin. The second group (E + SNP) received the same dose of endotoxin as well as a continuous infusion of sodium nitroprusside (SNP) to reduce PAP. Three hours after the endotoxin infusions, the sheep were terminated and the extravascular fluid-to-blood-free dry weight ratios of the lungs were determined (EVF). The base-line PAP was 17.5 +/- 2.7 mmHg. A two-way analysis of variance demonstrated a significant difference (P less than 0.01) in PAP between the E and E + SNP groups. Although PAP in each group varied as a function of time, the difference between the two groups did not. The mean PAP for the E + SNP group (20.9 +/- 1.5 mmHg) was lower than the E group PAP of 27.3 +/- 2.1 mmHg after the endotoxin spike. Furthermore, the E + SNP group EVF (3.9 +/- 0.2) was significantly less than the EVF of the E group (4.7 +/- 0.5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Regulation of increase in anastomotic blood flow following pulmonary arterial occlusion

We have previously shown that there is an acute increase in anastomotic bronchial blood flow (Qbr) after pulmonary arterial obstruction in dogs. We examined the role of arachidonic acid metabolites in mediating this increase. The left lower lobe (LLL) was isolated and perfused (zone 2) with autologous blood in open-chested anesthetized dogs (n = 19). Qbr was measured from the amount of blood that overflowed from the closed vascular circuit of the suspended LLL and changes in its weight. In the control animals, there was a prompt and significant increase in Qbr following pulmonary arterial obstruction. Pretreatment with indomethacin (n = 6) or sodium salicylate (n = 4) almost completely blocked this rise in Qbr. Following pulmonary arterial occlusion, there was a rise in both thromboxane and a prostacyclin metabolite (6-keto-PGF1 alpha) in the blood of the pulmonary circulation of the LLL, although the 6-keto-PGF1 alpha rose relatively more. Pretreatment with indomethacin caused a fall in both thromboxane and prostacyclin levels (n = 3), which no longer rose after pulmonary arterial occlusion. These findings suggested that the balance of the vasodilator (prostacyclin) and vasoconstrictor (thromboxane) prostaglandins may play an important role in mediating the rise in Qbr that follows pulmonary arterial obstruction.     

Histamine-induced changes in rat tracheal goblet cell mucin store and mucosal edema

The pathology of chronic asthma in human and mouse is characterized by inflammation and remodeling of airway tissues. As a result of repeated inflammatory insults to the lower airways, smooth muscle thickening, mucin secretion and airway hyperreactivity may develop. In ovalbumin (OVA)-sensitized mice with repeated challenges with OVA to the lower airways, the trachea and bronchi are characterized by goblet cell hyperplasia and mucus hypersecretion from goblet cells. Previous study reports that intravenous (i.v.) application of a high dose of capsaicin releases tachykinin from capsaicin-sensitive nerves, producing acute plasma leakage and mucosal edema formation and causing depletion of mucin granules in goblet cells that results in a reduction in the number and size of Alcian blue (AB)-positive goblet cells in the rat trachea within a few minute after capsaicin application. Histamine is an important non-neural mediator of asthma from mast cells. The present study investigated whether i.v. application of a high dose of histamine (18 μmol/ml/kg) could result in these acute changes and the similar time-course changes in rat trachea. The tracheal whole mounts stained with chloroacetate esterase reagent and AB and tracheal methacrylate sections stained with AB and periodic acid-Schiff reagent were used for evaluation of histological and cellular changes. At 5 min after histamine application, mucosal leaky venules were numerous and subepithelial edema ratio (% of length of edema along the mucosal epithelial circumference of tracheal cross section) was found to be 48.2 ± 4.9, which was greater (P < 0.01) than saline-treated rats. But, the number of AB-positive goblet cells, 2,030 ± 170/mm² of mucosal surface epithelium, was similar to saline-treated group (P > 0.05). One day later, edema ratio remained large and the number of AB-positive goblet cells was 1,140 ± 150/mm² epithelium, reduced to half the number of the group at 5 min after histamine (P < 0.01). It is suggested that mucus hypersecretion occurred at this time point. At 3 or 5 days after histamine, edema ratio gradually decreased. The number of AB-positive goblet cells continued to remain small on day 3. On day 5 after histamine, the number of AB-positive goblet cells restored to the level of rat group at 5 min after histamine application. At 7 days after histamine, edema ratio returned to the level of saline-treated group. It is concluded that degranulation and thinning of tracheal goblet cells and mucus hypersecretion lagged behind histamine-induced acute plasma leakage and edema, and restoration of mucin store in goblet cells was associated with remission of mucosal edema.     

A method for analysis of pulmonary arterial and venous occlusion data.

Recently, we presented a compartmental model of the pulmonary vascular resistance (R) and compliance (C) distribution with the configuration C1R1C2R2C3 (J. Appl. Physiol. 70: 2126-2136, 1991). This model was used to interpret the pressure vs. time data obtained after the sudden occlusion of the arterial inflow (AO), venous outflow (VO), or both inflow and outflow (DO) from an isolated dog lung lobe. In the present study, we present a new approach to the data analysis in terms of this model that is relatively simple to carry out and more robust. The data used to estimate the R's and C's are the steady-state arterial [Pa(0)] and venous [Pv(0)] pressures, the flow rate (Q), the area (A2) encompassed by Pa(t) after AO and the equilibrium pressure (Pd) after DO, and the average slope (m) of the Pa(t) and Pv(t) curves after VO. The following formulas can then be used to calculate the 2 R's and 3 C's: [Pa(0) - Pv(0)]/Q = R1 + R2 = RT, R1C1 congruent to to A2/[Pa(0) - Pd], R1 congruent to [Pa(0) - Pd]/Q, Q/m = C1 + C2 + C3 = CT, and C2 = CT - (RTC1/R2).     

Analysis of pulmonary arterial pressure profile after occlusion of pulsatile blood flow

In isolated canine lung lobes perfused with a pulsatile pump, arterial occlusions were performed and the postocclusion arterial pressure profiles were analyzed to estimate the pulmonary capillary pressure. A solenoid valve interposed between the pump and the lobar artery was used to perform arterial occlusions at several instants equally distributed within a pressure cycle. Double occlusions were also accomplished by simultaneously activating the solenoid valve and clamping the venous outflow of the lung lobe. To analyze an arterial occlusion pressure profile, we computed the best monoexponential fit of the pressure decay over a short period of time after the occlusion maneuvers. Two estimates of the capillary pressure were derived from this analysis: 1) the extrapolation of the exponential fit to the instant of occlusion, and 2) the point at which the recorded pressure decay curve merges with the exponential fit. The pressures thus determined were compared with the double occlusion pressure that provided an independent estimate of the pulmonary capillary pressure. Our results show that, under a wide range of conditions, the estimates of the capillary pressure obtained from the arterial occlusion data are nearly equal to the double occlusion pressures. Additionally, we estimated the capillary pressure variations within a pressure cycle by examining the occlusion pressures sampled at different instants of the cycle. The pulsatility of the pulmonary microvascular pressure varied with the pump frequency as well as the state of arterial and venous vasoaction. These variations are consistent with the representation of the lung vasculature as a low-pass filter.     

Isolation,culture and identification of pulmonary arterial smooth muscle cells from rat distal pulmonary arteries

The culture of pulmonary arterial smooth muscle cells (PASMCs) is one of the most powerful tools for exploring the mechanisms of pulmonary hypertension (PH). Both pulmonary vasoconstriction and remodeling occur predominantly in distal pulmonary arteries (PA). In this study, we provide our detailed and standardized protocol for easy isolation and culture of PASMCs from rat distal PA to supply every investigator with a simple, economical and useful method in studying PH. The protocol can be divided into four stages: isolation of distal PA, isolation of cells, growth in culture and passage of cells. Rat distal PASMCs were characterized by morphological activity and by immunostaining for smooth muscle α-actin and smooth muscle myosin heavy chain, but not for CD90/Thy-1 or von Willebrand factor. Furthermore, functional assessments were performed, confirming the presence of voltage-dependent Ca²⁺ channels and physiological characteristic of response to hypoxia. In conclusion, we have developed a detailed and simple protocol for obtaining rat distal PASMCs. These PASMCs exhibit features consistent with vascular smooth muscle cells, and they could subsequently be used to further explore the pathophysiological mechanisms of PH.     

Chrysin ameliorates ANTU‐induced pulmonary edema and pulmonary arterial hypertension via modulation of VEGF and eNOs

Alpha‐naphthylthiourea (ANTU), a rodenticide induces lung toxicity. Chrysin a flavonoid possesses antioxidant, anti‐inflammatory, and antihypertensive potential. The aim of this study was to evaluate the efficacy of chrysin against ANTU‐induced pulmonary edema (PE) and pulmonary arterial hypertension (PAH) in laboratory rats. Sprague‐Dawley rats were used to induce PE (ANTU, 10 mg/kg, ip) and PAH (ANTU, 5 mg/kg, ip, 4 weeks). Animals were treated with chrysin (10, 20, and 40 mg/kg) and various biochemical, molecular, and histological parameters were evaluated. Acute administration of ANTU induces PE revealed by significant (P < 0.05) increase in relative lung weight, pleural effusion volume, lung edema, bronchoalveolar lavage fluid cell counts, total protein, 5‐hydroxytryptamine (5‐HT), lactate dehydrogenase (LDH), and γ‐glutamyl transferase (GGT), whereas pretreatment with chrysin (20 and 40 mg/kg, ip) significantly (P < 0.05) attenuated these ANTU‐induced biochemical and histological alterations. Repeated administration of ANTU caused induction of PAH evaluated by significant (P < 0.05) alterations in electrocardiographic, hemodynamic changes, and left ventricular function, whereas chrysin (20 and 40 mg/kg, p.o.) treatment significantly (P < 0.05) attenuated these alterations. ANTU‐induced hematological and serum biochemical (aspartate transaminase, alanine transaminase, LDH, and creatinine kinase MB) alterations were significantly (P < 0.05) inhibited by chrysin. It also significantly (P < 0.05) decreased elevated levels of oxido‐nitrosative stress in the right ventricle (RV) and lung. Chrysin significantly (P < 0.05) attenuated downregulated endothelial nitric oxide synthase and upregulated vascular endothelial growth factor messenger RNA and protein expressions both in the RV and pulmonary artery. Chrysin inhibited ANTU‐induced PE and PAH via modulation of inflammatory responses (5‐HT, LDH, and GGT), oxido‐nitrosative stress, and VEGF and eNOs levels.