RESUMEN
No abstract available.
Asunto(s)
Neoplasias de la Mama/diagnóstico , Biopsia del Ganglio Linfático Centinela , Axila , Competencia Clínica , Reacciones Falso Negativas , Femenino , Humanos , Ganglios Linfáticos , Metástasis LinfáticaRESUMEN
Hyaluronan (HA) retention inside the synovial cavity of joints serves diverse protective roles. We tested the hypothesis that HA retention is mediated by the network of extracellular matrix proteins in the synovial lining. Cannulated rabbit knee joints were infused with HA solution with or without pretreatment by chymopapain, a collagen-sparing protease. Trans-synovial fluid escape rate was measured and, after a period of trans-synovial filtration, samples of intra-articular fluid and subsynovial fluid were analysed for HA to assess its trans-synovial ultrafiltration. In control joints, HA ultrafiltration was confirmed by postfiltration increases in intra-articular HA concentration (259 +/- 17% of infused concentration) and reduced subsynovial concentration (30 +/- 8%; n = 11). The proportion of HA molecules reflected by the synovium was 57-75%. Chymopapain treatment increased the hydraulic permeability of the synovial lining approximately 13-fold, almost abolished the trans-synovial difference in HA concentration and reduced the HA reflected fraction to 3-7% (n = 6; P < 0.001, ANOVA). Structural studies confirmed that chymopapain treatment depleted the matrix of proteoglycans but preserved its collagen. The findings thus demonstrate that HA ultrafiltration and synovial hydraulic permeability are determined by the network of non-collagen, extracellular matrix proteins. This may be important clinically, since protease activity is raised in rheumatoid arthritis, as are HA and fluid escape.
Asunto(s)
Quimopapaína/farmacología , Proteínas de la Matriz Extracelular/metabolismo , Ácido Hialurónico/metabolismo , Articulaciones/metabolismo , Líquido Sinovial/metabolismo , Animales , Pollos , Quimopapaína/administración & dosificación , Dextranos , Filtración , Fluoresceínas , Colorantes Fluorescentes , Inyecciones Intraarteriales , Articulaciones/efectos de los fármacos , Articulaciones/ultraestructura , Linfa/metabolismo , Conejos , Líquido Sinovial/efectos de los fármacos , Membrana Sinovial/efectos de los fármacos , Membrana Sinovial/ultraestructuraRESUMEN
OBJECTIVE: Hyaluronan (HA) has a major role in regulating synovial fluid volume. This role depends on the synovium functioning as an ultrafilter that reflects HA during trans-synovial fluid drainage. Reflection boosts the HA concentration on the membrane surface, leading to osmotic retention of synovial fluid ("buffering"). In arthritic effusions, however, HA concentration and osmotic buffering are greatly reduced. We tested the hypothesis that reflection is reduced (escape increased) when the HA concentration falls below the molecular entanglement concentration (C*). METHODS: HA at 0.2 mg/ml (
Asunto(s)
Ácido Hialurónico/metabolismo , Líquido Sinovial/metabolismo , Membrana Sinovial/metabolismo , Animales , Difusión , ConejosRESUMEN
In joint fluid the polymer hyaluronan (HA) confers viscous lubrication and greatly attenuates trans-synovial fluid loss (outflow buffering). Outflow buffering arises from the molecular sieving (reflection) and concentration polarization of HA at the synovial membrane surface. Outflow buffering declines if HA chain length is reduced, as in arthritis, and this has been attributed to reduced HA reflection. This was tested directly in the present study. Infused solutions of HA of approximately 2200 kDa (HA2000, 0.2 mg ml(-1)) or approximately 500 kDa (HA500, 0.2 mg ml(-1)) or approximately 140 kDa (HA140, 0.2-4.0 mg ml(-1)) were filtered across the synovial lining of the knee joint cavity of anaesthetized rabbits at a constant rate, along with a freely permeating reference solute, 20 kDa fluorescein-dextran (FD20). After a priming period the femoral lymph was sampled over 3 h. Mixed intra-articular (i.a.) fluid and subsynovial fluid were sampled at the end. Fluids were analysed by gel exclusion chromatography. The trans-synovial concentration profile was found to depend on polymer size. The i.a. concentration of HA2000 increased substantially relative to infusate and the subsynovial and lymph concentrations fell substantially. For HA500 and HA140 the trans-synovial concentration gradients were less pronounced, and absent for FD. The reflected fractions for HA2000, HA500 and HA140 across the cavity-to-lymph barrier were 0.65 +/- 0.05 (n = 10), 0.43 +/- 0.09 (n = 3) and 0.19 +/- 0.05 (n = 7), respectively, at matched filtration rates (P < 0.0001, analysis of variance). Reflected fractions calculated from HA i.a. accumulation or subsynovial dilution showed the same trend. The results demonstrate size-selective molecular sieving by the synovial extracellular matrix, equivalent to steric exclusion from cylindrical pores of radius 33-59 nm. The findings underpin the concentration polarization-outflow buffering theory and indicate that reduced HA chain length in arthritis exacerbates lubricant loss from a joint.
Asunto(s)
Matriz Extracelular/química , Ácido Hialurónico/química , Ácido Hialurónico/aislamiento & purificación , Articulación de la Rodilla/química , Líquido Sinovial/química , Membrana Sinovial/química , Ultrafiltración/métodos , Animales , Difusión , Conformación Molecular , Peso Molecular , Tamaño de la Partícula , Conejos , ViscosidadRESUMEN
Hyaluronan (HA), a component of synovial fluid, buffers fluid loss from joints. To explain this, a quantitative theory for HA concentration polarisation at a partially sieving synovial lining was developed. The theory predicts a fall in HA reflected fraction R with increased filtration rate. To test this, knees of anaesthetised rabbits were infused with HA and fluorescein-dextran (FD) at constant trans-synovial filtration rates of 6-89 microl min(-1). Samples of femoral lymph, mixed intra-articular fluid and subsynovial fluid after >/= 3 h were analysed by high-performance liquid chromatography. R was calculated as (1 - downstream/upstream concentration), using [FD] to adjust for joint lymph dilution in femoral lymph. Intra-articular HA concentration after >/= 3 h, 0.47 +/- 0.02 mg ml(-1) (mean +/-s.e.m., n= 31), exceeded the infusate concentration, 0.20 mg ml(-1), while subsynovial and lymph [HA] were reduced relative to [FD]. The changes in [HA] demonstrated synovial molecular sieving of HA. R from cavity to lymph (R(lymph)) fell monotonically from 0.93 at 6 microl min(-1) to 0.14 at 89 microl min(-1) (P < 0.0001, regression analysis, n= 33). R values calculated from the intra-articular HA accumulation (R(asp)) or the low subsynovial concentrations (R(syn)) were similar negative functions of filtration rate. R for lymphatic capillary endothelium (R(endo)), calculated from lymph/subsynovial concentration ratios, was effectively zero (-0.03 +/- 0.18, n= 21), confirming that synovium, not initial lymphatic endothelium, is the reflection site. Logarithmic linearisation of the results evaluated the synovial HA reflection coefficient as 0.91. In conclusion, the existence of concentration polarisation during joint fluid drainage was supported by the demonstration of a negative relation between filtration rate and R(lymph), R(asp) and R(syn).
Asunto(s)
Ácido Hialurónico/metabolismo , Articulaciones/metabolismo , Linfa/metabolismo , Algoritmos , Animales , Capilares/metabolismo , Pollos , Cromatografía Líquida de Alta Presión , Células Endoteliales/metabolismo , Filtración , Ácido Hialurónico/química , Técnicas In Vitro , Articulaciones/química , Cinética , Modelos Lineales , Linfa/química , Conejos , Líquido Sinovial/químicaRESUMEN
Synovial fluid hyaluronan (HA) profoundly buffers fluid loss from joints. This is attributed to the osmotic pressure of HA reflected by the joint lining. The aims were to quantify HA sieving during fluid drainage from joint to lymphatics and to compare the contributions of the synovial lining and lymphatic endothelium to sieving. HA (2100 kDa) and fluorescein-dextran (FD, 20 kDa) were infused under constant pressure into the knee cavity in anaesthetised rabbits. Samples were taken of femoral lymph and, after approximately 3-h transsynovial filtration, subsynovial fluid and mixed intra-articular fluid. [HA] and [FD] were analysed by HPLC and reflection was calculated as one minus transmitted fraction. Subsynovial and lymph [HA] were 16 and 12% of intra-articular [HA], which increased to 2.6 times infusate [HA] (P < 0.001, ANOVA, n = 19). [FD] was not significantly changed in infusate, aspirate, and subsynovial fluid but fell to 62% in femoral lymph due to dilution by skin/muscle lymph. The HA reflected fraction for the cavity-to-lymph barrier, R(lymph), was 0.54 +/- 0.03 (n = 82, mean +/- SEM), compared with 0.51 +/- 0.07 for cavity-to-subsynovium (R(syn), P > 0.05, Bonferroni) and 0.07 +/- 0.18 for subynovium-to-lymph (R(endo), P < 0.0001, Bonferroni). Lymphatic capillary endothelial reflection R(endo) was not significantly different from zero (one-sample t test). It is concluded that HA is partially sieved out of fluid leaving the joint cavity, and the sieve is the synovial lining interstitial matrix, not lymphatic capillary endothelium.
Asunto(s)
Endotelio Linfático/patología , Ácido Hialurónico/farmacología , Linfa/metabolismo , Vasos Linfáticos/patología , Membrana Sinovial/patología , Animales , Cromatografía Líquida de Alta Presión , Dextranos/farmacología , Endocitosis , Fémur/patología , Fluoresceínas/farmacología , Articulación de la Rodilla/patología , Modelos Teóricos , Presión Osmótica , Conejos , Membrana Sinovial/efectos de los fármacosRESUMEN
The coupling of periarticular lymph flow to transsynovial flow from a joint cavity, i.e., fluid load, is essential to avoid periarticular edema, which is associated with arthritic morning stiffness. To study coupling in swollen joints, a new method, "inside-out" cannulation, which eliminates dead space, resistance and cutout, was used to collect lymph from fine femoral lymph trunks in anesthetised rabbits while the knee joint cavity was infused with Evans blue albumin (EVA) at controlled intraarticular pressure and transsynovial drainage rates. The amount of joint lymph in femoral lymph (volume fraction V(v)) was calculated by EVA analysis. Joint lymph flow and EVA clearance was 1.5 +/- 0.4 microl min(-1) (mean +/- SEM, n = 62) at mean trans-synovial flow, 23 microl min(-1), and increased with pressure. Volume fraction increased from 16% at 10 cmH(2)O to 43% at 41 cmH(2)O. The increase in lymph flow with pressure, 0.052 +/- 0.025 microl min(-1) cmH(2)O(-1) (n = 61) was much smaller than the increase in transsynovial flow (periarticular fluid load) with pressure, 0.71 +/- 0.14 microl min(-1) cmH(2)O(-1) (P < 0.001). Their ratio, the coupling coefficient, was only 0.06-0.11. Thus, although up to 43% of femoral lymph could stem from a single swollen joint, this drained away only a small fraction of the transsynovial filtrate. The study showed that joint lymphatic drainage is coupled to joint pressure and transsynovial flow; but the coupling is insufficient to prevent periarticular fluid accumulation under conditions of joint volume expansion and limb immobility. This may contribute to the periarticular edema of arthritis.
Asunto(s)
Articulación de la Rodilla/metabolismo , Albúminas/metabolismo , Animales , Artritis/patología , Modelos Animales de Enfermedad , Linfa/metabolismo , Sistema Linfático/cirugía , Microcirculación/fisiología , Modelos Anatómicos , Presión , Conejos , Colorantes de Rosanilina/farmacología , Sinovectomía , Líquido Sinovial/fisiología , Membrana Sinovial/irrigación sanguínea , Factores de TiempoRESUMEN
The polysaccharide hyaluronan (HA) conserves synovial fluid by keeping outflow low and almost constant over a wide pressure range ('buffering'), but only at concentrations associated with polymer domain overlap. We therefore tested whether polymer interactions can cause buffering, using HA-chondroitin sulphate C (CSC) mixtures. Also, since it has been found that capillary filtration is insensitive to the Starling force interstitial osmotic pressure in frog mesenteries, this was assessed in synovium. Hyaluronan at non-buffering concentrations (0.50-0.75 mg ml(-1)) and/or 25 mg ml(-1) CSC (osmotic pressure 68 cmH(2)O) was infused into knees of anaesthetised rabbits in vivo. Viscometry and chromatography confirmed that HA interacts with CSC. Pressure (P(j)) versus trans-synovial flow (;Q(s)) relations were measured.;Q(s) was outwards for HA alone (1.2 +/- 0.9 microl min(-1) at 3 cmH(2)O, mean +/- S.E.M.; n = 6). CSC diffused into synovium and changed;Q(s) to filtration at low P(j) (-4.1 microl min(-1), 3 cmH(2)O, n = 5, P < 0.02, t test). Filtration ceased upon circulatory arrest (n = 3). At higher P(j), 0.75 mg ml(-1) HA plus CSC buffered;Q(s) to approximately 3 microl min(-1) over a wide range of P(j), with an outflow increase of only 0.04 +/- 0.02 microl min(-1) cmH(2)O(-1) (n = 4). With HA or CSC alone, buffering was absent (slopes 0.57 +/- 0.04 microl min(-1) cmH(2)O(-1) (n = 4) and 0.86 +/- 0.05 microl min(-1) cmH(2)O(-1) (n = 5), respectively). Therefore, polymer interactions can cause outflow buffering in joints. Also, interstitial osmotic pressure promoted filtration in fenestrated synovial capillaries, so the results for frog mesentery capillaries cannot be generalised. The difference is attributed to differences in pore ultrastructure.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Sulfatos de Condroitina/farmacología , Ácido Hialurónico/farmacología , Líquido Sinovial/metabolismo , Membrana Sinovial/metabolismo , Animales , Transporte Biológico/efectos de los fármacos , Permeabilidad Capilar/efectos de los fármacos , Interacciones Farmacológicas , Articulación de la Rodilla/metabolismo , Modelos Biológicos , Presión Osmótica , Polímeros/metabolismo , Conejos , Membrana Sinovial/irrigación sanguínea , Membrana Sinovial/efectos de los fármacos , ViscosidadRESUMEN
One month old soybean (Glycine max (L.) Merrill) cv. 'Williams' plants were exposed to nitrogen dioxide (NO2 at 0.1, 0.2, 0.3, and 0.5 microl liter(-1) and carbon filtered air (control), 7 h per day for five days, under controlled environment. Data were collected on net photosynthetic rate (PN), stomatal resistance (SR), and dark respiration rate (DR), immediately following the fifth day of exposure and 24 h after termination of exposure. Chlorophyll a (Ch a), chlorophyll b (Ch b), total chlorophyll (tot Ch) and foliar nitrogen (N) were measured before and after exposures. Growth characteristics--relative growth rate (RGR), net assimilation rate (NAR), leaf area ratio (LAR), and root shoot ratio (RSR) -- were computed for treated plants using standard growth equations. Increases of 18% and 23% in PN were observed immediately following exposure to 0.2 microl liter(-1) NO2 and after 24 h recovery period, respectively. With 0.5 microl liter(-1) NO2 treatment, reductions in PN of 23% and 50% were observed, immediately after exposure and following 24 h recovery, respectively. DR rates with 0.2 l liter(-1) treatment were higher than the control. Chlorophyll a and tot Ch showed significant reduction with 0.5 microl liter(-1) NO2 treatment. The percent reduction in Ch a and tot Ch with 0.5 microl liter(-1) NO2 were 45% and 47%, respectively. Increases in foliar nitrogen content after 0.2 and 0.3 microl liter(-1) NO2 treatments were 46% and 69%, respectively. Nitrogen dioxide at 0.5 microl liter(-1) reduced RGR and NAR by 47% and 51%, respectively. Leaf area ratio was 42% higher in 0.5 microl liter(-)1 NO2 treated plants, compared with the control; this increase was insufficient to compensate for the decrease in NAR resulting in a net decline in RGR. Nitrogen dioxide up to 0.2 microl liter(-1) increased PN and foliar-N content of soybean. With 0.5 microl liter(-1) NO2, significant decreases were observed in PN, leaf chlorophyll, foliar-N, NAR and RGR. Nitrogen dioxide up to 0.2 microl liter(-)1 has a favorable influence on overall growth characteristics of soybean; however, inhibitory effects were seen with NO2 treatment at 0.5 microl liter(-1).
RESUMEN
One-month-old soybean (Glycine max [L.] Merrill), cultivar 'Williams', plants were exposed to nitrogen dioxide (0.1, 0.2, 0.3 and 0.5 ppm) and carbon filtered air (control), 7 h per day, for 5 days, under a controlled environment. Leaf chlorophyll content (Ch a, Ch b, and total Ch content) and foliar nitrogen content (%N) were determined before and after the exposure. The influence of NO(2) treatments up to 0.3 ppm on leaf chlorophyll content was negligible although a stimulatory effect was evident in Ch a and total Ch content with 0.2 ppm NO(2). Marked decline in Ch content was observed with 0.5 ppm treatment; the reductions in Ch a and total Ch were 45% and 47%, respectively. Foliar-N contents of plants treated with 0.2 and 0.3 ppm NO(2) were higher than the control; plants exposed to 0.5 ppm NO(2) showed a 41% reduction in foliar-N compared to pre-exposure values.