Effect of longstanding venous stasis and hypoproteinaemia on lymph flow in the rat tail

This study was performed to provide information on the determinants of lymph flow by comparing the effect of venous stasis and hypoproteinaemia in the rat tail. This low-compliant tissue was chosen in an attempt to induce preferential changes in interstitial pressure or volume. The removal rate (kAlb) of 125I-labelled human serum albumin (I-HSA) injected subcutaneously was monitored with external gamma-counting equipment and used as a measure of lymph flow. Interstitial fluid hydrostatic pressure (Pi) was measured with wick-in-needle technique, and interstitial fluid was collected post mortem by dry wicks. Colloid osmotic pressure of plasma (COPp) and wick fluid (COPi) was measured with a colloid osmometer. In a separate group of experiments, 51Cr-EDTA and [125I]HSA were used to measure the interstitial fluid volume. Venous stasis, induced by bilateral ligation of the external tail veins, increased interstitial fluid hydrostatic pressure from 1.7 to 16 mmHg and kAlb from 0.030 to 0.063 h-1, whereas tail circumference was nearly constant. Interstitial volume averaged 1.17 ml/g dry weight in control animals and 1.27 ml/g during increased venous pressure. Daily injections of aminonucleoside in salt-loaded rats (0.3% NaCl as drinking water) reduced colloid osmotic pressure of plasma from 19.1 to 8.5 mmHg and of wick fluid from 11.2 to 2.9 mmHg, while interstitial fluid hydrostatic pressure increased to 5.2 mmHg. The removal rate of 125I-labelled human serum albumin increased to 0.113 h-1, compared to 0.051 h-1 in salt-loaded controls. The interstitial volume showed a marked increase in salt-loaded hypoproteinaemic rats, 1.75 ml/g dry weight, compared to 1.30 ml/g in salt-loaded controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

High vascular pressure-induced lung injury requires P450 epoxygenase-dependent activation of TRPV4

High vascular pressure targets the lung septal network, causing acute lung injury. While calcium entry in septal endothelium has been implicated, the channel involved is not known. This study tested the hypothesis that the vanilloid transient receptor potential channel, TRPV4, is a critical participant in the permeability response to high vascular pressure. Isolated lungs from TRPV4(+/+) or TRPV4(-/-) mice were studied at baseline or during high pressure challenge. Permeability was assessed via the filtration coefficient. Endothelial calcium transients were assessed using epifluorescence microscopy of the lung subpleural network. Light microscopy and point counting were used to determine the alveolar fluid volume fraction, a measure of alveolar flooding. Baseline permeability, calcium intensity, and alveolar flooding were no different in TRPV4(+/+) versus TRPV4(-/-) lungs. In TRPV4(+/+) lungs, the high pressure-induced permeability response was significantly attenuated by low calcium perfusate, the TRPV antagonist ruthenium red, the phospholipase A(2) inhibitor methyl arachidonyl fluorophosphonate, or the P450 epoxygenase inhibitor propargyloxyphenyl hexanoic acid. Similarly, the high pressure-induced calcium transient in TRPV4(+/+) lungs was attenuated with ruthenium red or the epoxygenase inhibitor. High vascular pressure increased the alveolar fluid volume fraction compared with control. In lungs from TRPV4(-/-) mice, permeability, calcium intensity, and alveolar fluid volume fraction were not increased. These data support a role for P450-derived epoxyeicosatrienoic acid-dependent regulation of calcium entry via TRPV4 in the permeability response to high vascular pressure.     

Capillary permeability to protein as a factor in the control of plasma volume

1. Measurements of plasma volume, blood volume, extracellular fluid volume and capillary permeability to albumin were made in rats before and after plasmapheresis extending over 2 days.2. In spite of a mean reduction of albumin concentration of nearly 40%, plasma volume showed a significant increase of 12%. There was a slight increase in blood volume and a slight decrease in extracellular fluid volume but these changes were not statistically significant.3. The mean capillary permeability to albumin was significantly lower after plasmapheresis than before and changes in permeability showed a strong negative correlation with changes in plasma volume. The possibility of a causal relationship is discussed.     

Determinants of peritubular capillary fluid uptake in hydropenia and saline and plasma expansion.

Hydrostatic (HPc) and oncotic (phic) pressures within the peritubular capillary, tubular pressure (Pt), nephron filtration rate, and plasma flow, and proximal fractional and absolute reabsorption (APR) were measured in anesthetized rats during hydropenia and plasma and saline expansion. Net interstitial pressure (phii-HPi) was estimated from subcapsular hydrostatic and oncotic pressures during saline expansion and these data were applied to a mathematical model of peritubular capillary fluid uptake to determine the profile of effective reabsorption pressure (ERP) with distance (x*) alongthe capillary and calculate the peritubular capillary permeability coefficient (LpAr). ERPX* = (PHIC MINUS HPC)X* MINUS (PHII MINUS HPi) and APR = ERPX*LpAr.During saline expansion phii minus HPi was -12.1 plus or minus 0.8 mmHg and ERP,3.8 mm. The LpAr was 0.07 nl/s per g KW per mmHg, and this value was applied to hypropenia and plasma expansion to determine ERP and phii minus HPi. The phiiminus HPi was +6.0 and +5.0 mmHg, respectively, and ERP was 4.1 and 3.5 mmHg.Efective reabsorptive pressure remained positive along x* in all states, and phii minus HPi correlated best changes in phic and poorly with changes in efferent plasma flow. The APR did not correlate with either calculated phii minus HPi or the transepithelial driving pressure, Pt + phii minus HPi.     

The pressure equilibrating function of pars flaccida in middle ear mechanics

In the adult male rat the precise volumes of the middle ear cavity, the lateral attic compartment and a maximally retracted pars flaccida were calculated to 550 microliter, 21 microliter and 3 microliter respectively, using Woods metal. Small intratympanal pressure or volume changes caused the pars flaccida to move in a medial-lateral direction. As the volume of a maximally retracted or bulging pars flaccida only amounted to about 0.5% of the total middle ear volume and the pars flaccida reacted even at volume changes of 0.1% of the middle ear, it would seem that the pressure equilibrating capacity of the elastic pars flaccida is negligible. On the other hand the pars flaccida is extremely sensitive to pressure and volume changes in the middle ear and may take part in the pressure equilibrating system as a detector for minute pressure and/or volume changes in the middle ear.     

Impact of supplemental oxygen in mechanically ventilated adult and infant mice

The aim of the present study was to determine the short-term effects of hyperoxia on respiratory mechanics in mechanically ventilated infant and adult mice. Eight and two week old BALB/c mice were exposed to inspired oxygen fractions [Formula: see text] of 0.21, 0.3, 0.6, and 1.0, respectively, during 120 min of mechanical ventilation. Respiratory system mechanics and inflammatory responses were measured. Using the low-frequency forced oscillation technique no differences were found in airway resistance between different [Formula: see text] groups when corrected for changes in gas viscosity. Coefficients of lung tissue damping and elastance were not different between groups and showed similar changes over time in both age groups. Inflammatory responses did not differ between groups at either age. Hyperoxia had no impact on respiratory mechanics during mechanical ventilation with low tidal volume and positive end-expiratory pressure. Hence, supplemental oxygen can safely be applied during short-term mechanical ventilation strategies in infant and adult mice.     

Otitis media in a mouse model for Down syndrome

The Ts65Dn mouse shares many phenotypic characteristics of human Down syndrome. Here, we report that otitis media, characterized by effusion in the middle ear and hearing loss, was prevalent in Ts65Dn mice. Of the 53 Ts65Dn mice tested, 81.1% had high auditory-evoked brainstem response (ABR) thresholds for at least one of the stimulus frequencies (click, 8 kHz, 16 kHz and 32 kHz), in at least one ear. The ABR thresholds were variable and showed no tendency toward increase with age, from 2 to 7 months of age. Observation of pathology in mice, aged 3–4 months, revealed middle ear effusion in 11 of 15 Ts65Dn mice examined, but only in two of 11 wild-type mice. The effusion in each mouse varied substantially in volume and inflammatory cell content. The middle ear mucosae were generally thickened and goblet cells were distributed with higher density in the epithelium of the middle ear cavity of Ts65Dn mice as compared with those of wild-type controls. Bacteria of pathogenic importance to humans also were identified in the Ts65Dn mice. This is the first report of otitis media in the Ts65Dn mouse as a model characteristic of human Down syndrome.     

Relationship among circulating leukocytes, platelets, and microvascular responses during induction of chronic colitis

The mechanisms by which microvascular alterations contribute to the pathogenesis of the inflammatory bowel diseases (IBDs; Crohn's disease, ulcerative colitis) have not been clearly delineated. The purpose of the current study was to characterize the inflammatory events, microvascular alterations, and blood cell changes that occur in a mouse model of IBD. In this model, CD4⁺ T-lymphocytes obtained from interleukin-10-deficient mice were injected intraperitoneally into lymphopenic, recombinase-activating gene-1 deficient (RAG^−/−) mice. Two groups of control mice were also included: RAG^−/− mice and C57BL/6 mice that were injected with phosphate-buffered saline but did not receive the T-cells. Four weeks later, the RAG^−/− mice that had received the T-cell transfer showed significant signs of colonic inflammation, but without significant decreases in either body weight or mean arterial blood pressure. T-cell transfer increased the volume % of circulating platelets, while decreasing the number of circulating red blood cells. Additionally, the T-cell transfer tended to increase the circulating numbers of both lymphocytes and neutrophils when compared to unmanipulated RAG^−/− mice. First-order colonic arterioles and venules tended to dilate in the colitic mice; however, the dilation was considerably more substantial with higher numbers of circulating leukocytes. The possibility that circulating inflammatory cells initiate the microvascular alterations in colitis warrants further investigation.     

"清气凉营"注射液对病毒和微循环的药理作用

目的 研究“清气凉营”对病毒和微循环的作用。方法 在MDCK，MESO细胞和小白鼠上进行体外和体内抗病毒实验。在家兔身上感染仙台病毒形成高热模型。静脉注射啤酒酵母形成家兔微循环障碍。应用Olympus-BH微循环显微系统，分别观察家兔、大白鼠和小白鼠的球结膜、肠系膜和耳壳微循环，应用MCIMPS-B图象分析仪测定微血管管径，应用Evens兰测定毛细血管通透性。结果 “清气凉营”对体外和体内出血热病毒和流感病毒有强烈抑制作用。对家兔高热有退热效应。扩张大鼠肠系膜和小鼠耳壳微动脉和微静脉，血液流速增快，流量增加。“清气凉营”能降低组织胺引起大鼠和小鼠血管通透性增加的作用，并使微循环障碍恢复到正常。结论“清气凉营”对体外和体内的出血热病毒和流感病毒有较强的抑制作用，增强机体免疫功能，对仙台病毒引起的高热模型有退热作用。“清气凉营”可扩张微血管，增快血液流速，增加流量，降低由组织胺引起的血管通透性增加的作用，改善微循环，促进康复。     

美声喉泰含片药效学研究

用小鼠建立咳嗽、炎症、疼痛等模型,观察其咳嗽次数、肿胀程度、毛细血管通透性、扭体次数等试验,结果表明美声喉泰含片对氨水引起的小鼠咳嗽有显著的镇咳作用,对醋酸刺激引起的小鼠腹腔毛细血管通透性增高有显著的抑制作用,对巴豆油引起小鼠耳肿胀有一定抑制作用,对醋酸引起的小鼠疼痛扭体反应有显著的抑制作用。     

Water permeability through aquaporin-4 is regulated by protein kinase C and becomes rate-limiting for glioma invasion

Glial-derived tumors, gliomas, are highly invasive cancers that invade normal brain through the extracellular space. To navigate the tortuous extracellular spaces, cells undergo dynamic changes in cell volume, which entails water flux across the membrane through aquaporins (AQPs). Two members of this family, AQP1 and AQP4 are highly expressed in primary brain tumor biopsies and both have a consensus phosphorylation site for protein kinase C (PKC), which is a known regulator of glioma cell invasion. AQP4 colocalizes with PKC to the leading edge of invading processes and clustered with chloride channel (ClC2) and K⁺–Cl⁻ cotransporter 1 (KCC1), believed to provide the pathways for Cl⁻ and K⁺ secretion to accomplish volume changes. Using D54MG glioma cells stably transfected with either AQP1 or AQP4, we show that PKC activity regulates water permeability through phosphorylation of AQP4. Activation of PKC with either phorbol 12-myristate 13-acetate or thrombin enhanced AQP4 phosphorylation, reduced water permeability and significantly decreased cell invasion. Conversely, inhibition of PKC activity with chelerythrine reduced AQP4 phosphorylation, enhanced water permeability and significantly enhanced tumor invasion. PKC regulation of AQP4 was lost after mutational inactivation of the consensus PKC phosphorylation site S180A. Interestingly, AQP1 expressing glioma cells, by contrast, were completely unaffected by changes in PKC activity. To demonstrate a role for AQPs in glioma invasion in vivo, cells selectively expressing AQP1, AQP4 or the mutated S180A-AQP4 were implanted intracranially into SCID mice. AQP4 expressing glioma cells showed significantly reduced invasion compared to AQP1 and S180 expressing tumors as determined by quantitative stereology, consistent with a differential role for AQP1 and AQP4 in this process.     

Unrestrained Acoustic Plethysmograph for Measuring Tidal Volume in Mice

The traditional method for measurement of tidal volume in unrestrained mice relies on pressure changes induced by a freely respiring animal in a whole body plethysmograph. These changes have been assumed to be the result of thermo-hygrometric differences between respired air and gas within the chamber. It is known, however, that gas compression in the lung can also contribute significantly to changes in plethysmograph pressure. This study describes an acoustic plethysmograph for mice that is capable of measuring the tidal volume time series without the errors associated with the traditional method.The plethysmograph was designed as a resonating cavity at a fixed frequency. It had a sharp resonant peak and was tuned so that changes in body volume produced nearly linear changes in sound amplitude. The plethysmograph was tested with a water filled balloon connected to a syringe pump. The volume of the balloon was varied as a triangle wave with an amplitude of 250 μL. The RMS error between measured and delivered volume was 4.43 μL. A volume step test, performed to assess the response time of the system, showed that the plethysmograph responded in less than one millisecond.     

Peripheral neuropathy in the Twitcher mutant. A new experimental model of endoneurial edema

The Twitcher mouse (Twi/Twi) is a recently identified mutant experimental model for human globoid leukodystrophy. Affected mice develop neurologic abnormalities with demyelination of white matter and peripheral nerve due to an inherited enzyme deficiency. The neuropathy has unusual pathologic features:severe interstitial edema and infiltration by eosinophils. To investigate its pathogenesis and to identify the mechanism of demyelination, we studied vascular permeability and measured endoneurial fluid pressure. Significantly increased endoneurial fluid pressure was detected in clinically affected animals (average, 6.4 cm H2O) versus controls (1.7 cm H2O), and these data are the first measurements of EFP to be reported in mice. Increased vascular permeability to horseradish peroxidase was visualized by electron microscopy with leakage of horseradish peroxidase between endothelial cells and flooding of the endoneurial interstitium. Numerous eosinophils were present in the interstitium, as well as some polymorphonuclear cells, occasional erythrocytes, and degranulating mast cells. Abnormalities of nerve fibers included swelling of Schwann cells with intracytoplasmic inclusions, demyelination, and remyelination. As well as being a model for globoid leukodystrophy, the Twitcher is the first spontaneously occurring experimental model for endoneurial edema and increased endoneurial fluid pressure.     

Atrial natriuretic peptide modulation of albumin clearance and contrast agent permeability in mouse skeletal muscle and skin: role in regulation of plasma volume

Atrial natriuretic peptide (ANP) via its guanylyl cyclase-A (GC-A) receptor participates in regulation of arterial blood pressure and vascular volume. Previous studies demonstrated that concerted renal diuretic/natriuretic and endothelial permeability effects of ANP cooperate in intravascular volume regulation. We show that the microvascular endothelial contribution to the hypovolaemic action of ANP can be measured by the magnitude of the ANP-induced increase in blood-to-tissue albumin transport, measured as plasma albumin clearance corrected for intravascular volume change, relative to the corresponding increase in ANP-induced renal water excretion. We used a two-tracer method with isotopically labelled albumin to measure clearances in skin and skeletal muscle of: (i) C57BL6 mice; (ii) mice with endothelium-restricted deletion of GC-A (floxed GC-A × tie2-Cre: endothelial cell (EC) GC-A knockout (KO)); and (iii) control littermates (floxed GC-A mice with normal GC-A expression levels). Comparison of albumin clearances in hypervolaemic EC GC-A KO mice with normovolaemic littermates demonstrated that skeletal muscle albumin clearance with ANP treatment accounts for at most 30% of whole body clearance required for ANP to regulate plasma volume. Skin microcirculation responded to ANP similarly. Measurements of permeability to a high molecular mass contrast agent (35 kD Gadomer) by dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) enabled repeated measures in individual animals and confirmed small increases in muscle and skin microvascular permeability after ANP. These quantitative methods will enable further evaluation of the contribution of ANP-dependent microvascular beds (such as gastro-intestinal tract) to plasma volume regulation.     

Alcohol consumption and blood-cerebrospinal fluid barrier dysfunction in man

The cerebrospinal fluid (CSF)/serum albumin ratio has been used a marker for blood CSF barrier permeability in 116 normal patients. We attempted to correlate the CSF/serum albumin ratio with a number of clinically measurable parameters including alcohol consumption. Alcohol consumption had a significant effect on the blood-CSF barrier. Our data indicate that alcohol increases blood CSF barrier permeability in a dose-dependent manner. The measured values of parameters indirectly indicative of alcohol consumption, such as gamma-glutamyltranspeptidase (gamma-GT) and erythrocyte mean corpuscular volume (MCV), were also correlated with enhanced blood-CSF barrier permeability. Although an apparent influence of age, body weight and sex on blood-CSF barrier permeability was observed, these correlations were not separable from the effect of alcohol consumption.     

Interstitial fluid flow in tendons or ligaments: A porous medium finite element simulation

The purpose of this study is to describe interstitial fluid flow in axisymmetric soft connective tissue (ligaments or tendons) when they are loaded in tension. Soft hydrated tissue was modelled as a porous medium (using Darcy’s Law), and the finite element method was used to solve the resulting equations governing fluid flow. A commercially available computer program (FiDAP) was used to create an axisymmetric model of a biomechanically tested rat ligament. The unknown variables at element nodes were pressure and velocity of the interstitial fluid (Newtonian and incompressible). The effect of variations in fluid viscosity and permeability of the solid matrix was parametrically explored. A transient loading state mimicking a rat ligament mechanical experiment was used in all simulations. The magnitude and distribution of pressure, stream lines, shear (stress) rate, vorticity and velocity showed regular patterns consistent with extension flow. Parametric changes of permeability and viscosity strongly affected fluid flow behaviour. When the radial permeability was 1000 times less than the axial permeability, shear rate and vorticity increased (approximately 5-fold). These effects (especially shear stress and pressure) suggested a strong interaction with the solid matrix. Computed levels of fluid flow suggested a possible load transduction mechanism for cells in the tissue.     

Inotropic and intracellular acid-base changes during metabolic acidosis.

Experiments in isolated, Ringer-perfused isovolumic rabbit hearts showed that metabolic acidosis resulted in a decrease in peak left ventricular pressure and dP/dt. Concomitantly, the decrease in extracellular pH from 7.28 plus or minus 0.02 to 6.82 plus or minus 0.02 at constant PaCO2 was associated with a negative av HCO3- difference that lasted throughout the duration of acidosis. This negative av HCO3- difference indicated that either HCO3- moved into the intravascular space or H+ moved in the opposite direction during acidosis. During perfusion with normal pH solution av HCO3- was not significantly different from zero. Washout of the extracellular space with the acid solution can account for only 32 percent of the total amount of HCO3-recovered in the venous perfusate during the 30 min of acidosis. The remaining 68 percent (10.7 plus or minus 3.1 mmol times kg-1 of cardiac tissue) must then have originated in the cellular fluid. When intracellular fluid volume is taken into account, it can be calculated that 21.3 plus or minus 6.1 mmol of HCO3- moved into the vascular space per liter of intracellular water. The magnitude of this HCO3- flux suggests that significant myocardial cellular acid-base changes are associated with metabolic acidosis.     

Lowered albumin extravasation rate in heart but not in other organs in β3‐integrin‐deficient mice

Aim: The vascular protein permeability is dependent on the integrity of the vascular wall. The heart capillaries in male mice lacking β3 integrins have an immature phenotype. Previously, we have demonstrated a role for αvβ3 integrins in control of interstitial fluid pressure (Pif) and thereby in the fluid flux during inflammation. We wanted to explore a possible role for αvβ3 integrins in controlling capillary protein permeability during control situation and inflammation. Methods: We performed double‐tracer and microdialysis experiments on β3‐integrin‐deficient mice and wild type control mice. We also measured blood pressure and heart rate in the two mice strains. Results: We found reduced albumin extravasation (during 25 min) in the heart capillaries (0.053 ± 0.003 vs. 0.087 ± 0.009 mL g^?1 dw, P < 0.05), and an increased cardiac mass/body weight (5.3 × 10^?3 ± 0.3 × 10^?3 vs. 3.8 × 10^?3 ± 0.1 × 10^?3, P < 0.01) in the β3‐integrin‐deficient mice (n = 6) compared with the controls (n = 6). Heart rate and blood pressure were the same in mice with and without β3‐integrins. No difference in permeability was found in other tissues studied, or under local inflammation. Conclusion: These results show a function for the αvβ3 integrin in the regulation of protein permeability, selective for the heart capillaries.     

Nrf2 is closely related to allergic airway inflammatory responses induced by low-dose diesel exhaust particles in mice

We have recently reported that disruption of nuclear erythroid 2 P45-related factor 2 (Nrf2) enhances susceptibility to airway inflammatory responses induced by low-dose diesel exhaust particles (DEP) in mice. C57BL/6 Nrf2 knockout (Nrf2^−/−) mice and wild-type (Nrf2^+/+) mice were further exposed to low-dose DEP for 7 h/day, 5 days/week, for a maximum of 8 weeks. After exposure to DEP for 5 weeks, allergic airway inflammation was generated in the mice by intraperitoneal sensitization with OVA followed by intranasal challenge. Nrf2^−/− mice exposed to relatively low-dose DEP showed significantly increased percentage changes relative to the OVA alone group in terms of airway hyperresponsiveness (AHR) and inflammatory cells, levels of IL-5 and thymus and activation regulated chemokine (TARC) in bronchoalveolar lavage (BAL) fluid than did Nrf2^+/+ mice. Lung tissues of Nrf2^−/− mice after DEP exposure showed inflammatory cell infiltrates, and increased PAS staining-positive mucus cell hyperplasia. In contrast, the percentage changes relative to the OVA group in the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio in whole blood was higher in Nrf2^+/+ mice than in Nrf2^−/− mice. By using Nrf2^−/− mice, it was shown for the first time that relatively low-dose DEP exposure induces oxidant stress, and that host anti-oxidant responses play a key role in the development of DEP-induced exacerbation of allergic airway inflammation.     

Oedema-preventing mechanisms in a low-compliant tissue: studies on the rat tail

The long-term effect of elevated tail venous pressure (Pv) on interstitial fluid pressure (Pi) and colloid osmotic pressure (COPi) was studied in rats. Measurements were performed on different rats after 1-5 days of congestion, and on a sham operated control group. Two different degrees of venous stasis were obtained by means of graded ligation of tail veins and skin. Moderate degree of ligation (group 1) caused no visual oedema. More extensive ligation (group 2) gave marked oedema already on day 1 and further progression and exudation from the skin during the following 3 days. Tail venous pressure was measured by micropuncture and interstitial fluid pressure by the wick-in-needle technique. Interstitial fluid was sampled by a dry wick method, and colloid osmotic pressure was measured with a membrane osmometer. In group 1, Pv had increased from 6.5 to 21.5 mmHg on day 1, and thereafter remained unchanged. Interstitial fluid pressure increased from 3.4 to 16.9 mmHg after ligation and to 19.2 mmHg on day 1. Colloid osmotic pressure fell from 11.7 to 5.8 mmHg. From day 1, both interstitial fluid pressure and colloid osmotic pressure remained largely unchanged. In group 2 rats, tail venous pressure increased to 33.0 and 36.3 mmHg on day 1 and 2 and then fell towards control level during the following 3 days. Interstitial fluid pressure increased to 22.9 and 31.4 mmHg before a rapid decline towards control level. Colloid osmotic pressure was reduced from 11.7 to 5.6 mmHg. We conclude that hydrostatic counterpressure may provide an oedema-preventing effect of up to about 15 mmHg.(ABSTRACT TRUNCATED AT 250 WORDS)