RBCs distribute oxygen to tissues, but, paradoxically, blood transfusion does not always improve oxygen delivery and is associated with ischemic events. We hypothesized that storage of blood would ...result in loss of NO bioactivity, impairing RBC vasodilation and thus compromising blood flow, and that repleting NO bioactivity would restore RBC function. We report that S-nitrosohemoglobin (SNO-Hb) concentrations declined rapidly after storage of fresh venous blood and that hypoxic vasodilation by banked RBCs correlated strongly with the amounts of SNO-Hb (r² = 0.90; P < 0.0005). Renitrosylation of banked blood during storage increased the SNO-Hb content and restored its vasodilatory activity. In addition, canine coronary blood flow was greater during infusion of renitrosylated RBCs than during infusion of S-nitrosothiol-depleted RBCs, and this difference in coronary flow was accentuated by hypoxemia (P < 0.001). Our findings indicate that NO bioactivity is depleted in banked blood, impairing the vasodilatory response to hypoxia, and they suggest that SNO-Hb repletion may improve transfusion efficacy.
In animals, the accepted model of carbohydrate digestion and absorption involves reduction of disaccharides into the monosaccharides glucose, fructose, and galactose followed by their individual ...transmembrane transport into cells. In 2011, a gene for a distinct disaccharide sucrose transporter (SCRT) was found in
Drosophila melanogaster
and characterized in a yeast expression system. The purpose of the present investigation was to functionally identify and characterize a putative disaccharide transporter analog in the hepatopancreas of the American lobster,
Homarus americanus
. Purified hepatopancreatic brush-border membrane vesicles (BBMV) were used in transport experiments using
14
C-sucrose and a Millipore filter isolation technique. In the absence of sodium, an external pH of 4 significantly stimulated the uptake of
14
C-sucrose compared to that occurring at pH 5, 6, or 7. At pH 7, increasing external concentrations of sodium increased
14
C-sucrose uptake by BBMV in a hyperbolic fashion and this stimulation was significantly reduced when the pH was changed to 4, suggesting that both protons and sodium ions were each capable of driving the uptake of the sugar. In experiments with a variety of monosaccharides, disaccharides, and trisaccharides, used as potential inhibitors of
14
C-sucrose uptake, only maltose and trehalose inhibited carrier-mediated
14
C-sucrose transport. An additional experiment showed that 20 mM maltose was a competitive inhibitor of
14
C-sucrose uptake. The use of a putative lobster SCRT by both maltose and trehalose is nutritionally appropriate for lobsters as they commonly digest glycogen and chitin, polymers of maltose and trehalose, respectively. These findings suggest there is a brush-border proton- or sodium-dependent, hepatopancreatic carrier process, shared by sucrose, maltose, and trehalose, that may function to absorb disaccharides that are produced from digestion of naturally occurring dietary constituents.
Pulmonary arterial hypertension (PAH) is often difficult to diagnose and challenging to treat. Untreated, it is characterized by a progressive increase in pulmonary vascular resistance leading to ...right ventricular failure and death. The past decade has seen remarkable improvements in therapy, driven largely by the conduct of randomized controlled trials. Still, the selection of most appropriate therapy is complex, and requires familiarity with the disease process, evidence from treatment trials, complicated drug delivery systems, dosing regimens, side effects, and complications. This chapter will provide evidence-based treatment recommendations for physicians involved in the care of these complex patients. Due to the complexity of the diagnostic evaluation required, and the treatment options available, it is strongly recommended that consideration be given to referral of patients with PAH to a specialized center.
Gill chambers of the Atlantic lobster,
Homarus americanus
, possess three structures that are involved with respiration and ion regulation: gill filaments, epipodites, and branchiostegites. This ...paper describes ion transport mechanisms present in the plasma membranes of branchiostegite epithelial cells and the effects of pH on the uptake of
45
Ca by these processes. Partially purified membrane vesicles (PPMV) of branchiostegite cells were produced by a homogenization/centrifugation method that has previously been used to define ion transport processes in both crab and lobster gill tissues. In the present study, lobster branchiostegite PPMV
45
Ca uptake was highest at pH 8.5 and lowest at pH values between 6.0 and 7.0 (
p
< 0.02). At pH 8.0,
45
Ca uptake was a biphasic process consisting of a saturable process at low Ca and a linear process at higher Ca. At pH 6.0,
45
Ca uptake was only a linear process and paralleled linear uptake at pH 8.0. A valinomycin/K
+
-induced membrane potential (PD, inside negative) doubled
45
Ca uptake at pH 7.0 above that in the absence of a PD (
p
< 0.05). An induced PD at pH 8.0 did not significantly (
p
> 0.05) affect
45
Ca uptake observed in the absence of a PD, but was threefold greater than uptake at pH 7.0 in the absence of a PD (
p
< 0.05). Amiloride (2 mM) did not affect
45
Ca uptake at pH 8.0, but 2 mM amiloride + 100 µM verapamil reduced uptake by approximately 50%. In the presence of both 2 mM amiloride + 100 µM verapamil, 15 s
45
Ca influx at pH 8.5 was a hyperbolic function of Ca (0.1–5 mM) (
K
m
= 4.2 ± 0.3 mM;
J
max
= 9792 ± 439 pmol/mg protein × 15 s).
45
Ca influxes at pH 7.5 under the same conditions were also hyperbolic with
K
m
= 8.3 ± 1.4 mM;
J
max
= 10732 ± 1250 pmol/mg protein × 15 s.
K
m
values were significantly different (
p
< 0.05), but
J
max
values were not (
p
> 0.05). These results suggest that
45
Ca uptake by lobster branchiostegites may have occurred by the combination of diffusion through a verapamil-inhibited calcium channel and carrier-mediated transport by amiloride-insensitive, electroneutral, 1Ca
2+
/2H
+
antiporters. Decreased pH, as might occur during ocean acidification, did not appear to modify calcium diffusion through the channels, but protons acted as competitive inhibitors of calcium transport by carrier-mediated antiport. Decreased calcium uptake with continued ocean acidification may significantly affect calcification processes during periodic molting, potentially influencing mortality.
Previous work with isolated outer membrane vesicles of lobster branchiostegite epithelial cells has shown that
45
Ca
2+
uptake by these structures is significantly (
p
< 0.02) reduced by an ...incremental decrease in saline pH (increased proton concentration) and that this decrease is due to competitive inhibition between carrier-mediated transport of
45
Ca
2+
and hydrogen ions. The present paper extends these previous findings and describes the combined effects of pH and cationic heavy metals on branchiostegite uptake of
45
Ca
2+
. Partially purified membrane vesicles of branchiostegite cells were produced by a homogenization/centrifugation method and were loaded with mannitol at pH 7.0. The time course of 1 mM
45
Ca
2+
uptake in a mannitol medium at pH 8.5 containing 100 µM verapamil (Ca
2+
channel blocker) was hyperbolic and approached equilibrium at 30 min. This uptake was either significantly reduced (
p
< 0.05) by the addition of 5 µM Zn
2+
or essentially abolished with the addition of 5 µM Cu
2+
. Increasing zinc concentrations (5–500 µM) reduced 1 mM
45
Ca
2+
uptake at pH 8.5 or 7.5 in a hyperbolic fashion with the remaining non-inhibited uptake due to apparent non-specific binding. Uptake of 1 mM
45
Ca
2+
at pH 8.5, 7.5, 7.5 + Zn
2+
, and 7.5 + Zn
2+
+ Cu
2+
+ Cd
2+
in the presence of 100 µM verapamil displayed a stepwise reduction of
45
Ca
2+
uptake with the addition of each treatment until only non-specific isotope binding occurred with all cation inhibitors.
45
Ca
2+
influxes (15 s uptakes; 0.25–5.0 mM calcium + 100 µM verapamil) in the presence and absence of 10 µM Zn
2+
were both hyperbolic functions of calcium concentration. The curve with Zn
2+
displayed a transport
K
m
twice that of the control (
p
< 0.05), while inhibitor and control curve
J
max
values were not significantly different (
p
> 0.05), suggesting competitive inhibition between
45
Ca
2+
and Zn
2+
influxes. Analysis of the relative inhibitory effects of increased proton or heavy metal interaction with
45
Ca
2+
uptake suggests that divalent metals may reduce the calcium transport about twice as much as a drop in pH, but together, they appear to abolish carrier-mediated transport.
Transepithelial absorption of dietary sucrose in the American lobster,
Homarus americanus
, was investigated by mounting an intestine in a perfusion chamber to characterize mucosal to serosal (MS)
14
...C-sucrose transport. These fluxes were measured by adding varying concentrations of
14
C-sucrose to the perfusate and monitoring their appearance in the bathing solution. Transepithelial
14
C-sucrose transport was the combination of a hyperbolic function of luminal concentration, following Michaelis–Menten kinetics, and apparent diffusion. The kinetic constants of the putative sucrose transporter were
K
M
= 20.50 ± 6.00 µM and
J
max
= 1.81 ± 0.50 pmol/cm
2
× min. Phloridzin, an inhibitor of Na
+
-dependent mucosal glucose transport, decreased MS
14
C-sucrose transport. Decreased MS
14
C-sucrose transport also occurred in the presence of luminal trehalose, a disaccharide containing
d
-glucose moieties. Thin-layer chromatography (TLC) identified the chemical nature of radioactively labeled sugars in the bath following transepithelial transport. TLC revealed
14
C-sucrose was transported across the intestine largely intact with no
14
C-glucose or
14
C-fructose appearing in the serosal bath or luminal perfusate. Only 13 % of bath radioactivity was volatile metabolites. Results suggest that disaccharide sugars can be transported intact across crustacean intestine and support the occurrence of a functional disaccharide membrane transporter.
Elasmobranchs are considered to be top marine predators, and in general play important roles in the transfer of energy within marine ecosystems. Despite this, little is known regarding the ...physiological processes of digestion and nutrient absorption in these fishes. One topic that is particularly understudied is the process of nutrient uptake across the elasmobranch gastrointestinal tract. Given their carnivorous diet, the present study sought to expand knowledge on dietary nutrient uptake in elasmobranchs by focusing on the uptake of products of protein digestion. To accomplish this, a full-length cDNA encoding peptide transporter 1 (PepT1), a protein previously identified within the brush border membrane of vertebrates that is responsible for the translocation of peptides released during digestion by luminal and membrane-bound proteases, was isolated from the bonnethead shark (
Sphyrna tiburo
). A cDNA encoding the related peptide transporter PepT2 was also isolated from
S. tiburo
using the same methodology. The presence of PepT1 was then localized in multiple components of the bonnethead digestive tract (esophagus, stomach, duodenum, intestine, rectum, and pancreas) using immunohistochemistry. Vesicle studies were used to identify the apparent affinity of PepT1 and to quantify the rate of dipeptide uptake by its H
+
-dependent cotransporter properties. The results of this study provide insight into the properties of peptide uptake within the bonnethead gut, and can facilitate future work on physiological regulation of protein metabolism and absorption including how these processes may vary in elasmobranchs that exhibit different feeding strategies.
Hepatopancreatic brush border membrane vesicles (BBMV), made from Atlantic White shrimp (
Litopenaeus setiferus
), were used to characterize the transport properties of
3
H-
l
-leucine influx by ...these membrane systems and how other essential amino acids and the cations, sodium and potassium, interact with this transport system.
3
H-
l
-leucine uptake by BBMV was pH-sensitive and occurred against transient transmembrane concentration gradients in both Na
+
- and K
+
-containing incubation media, suggesting that either cation was capable of providing a driving force for amino acid accumulation.
3
H-
l
-leucine uptake in NaCl or KCl media were each three times greater in acidic pH (pH 5.5) than in alkaline pH (pH 8.5). The essential amino acid,
l
-methionine, at 20 mM significantly (
p
< 0.0001) inhibited the 2-min uptakes of 1 mM
3
H-
l
-leucine in both Na
+
- and K
+
-containing incubation media. The residual
3
H-
l
-leucine uptake in the two media were significantly greater than zero (
p
< 0.001), but not significantly different from each other (
p
> 0.05) and may represent an
l
-methionine- and cation-independent transport system.
3
H-
l
-leucine influxes in both NaCl and KCl incubation media were hyperbolic functions of
l
-leucine, following the carrier-mediated Michaelis–Menten equation. In NaCl,
3
H-
l
-leucine influx displayed a low apparent
K
M
(high affinity) and low apparent
J
max
, while in KCl the transport exhibited a high apparent
K
M
(low affinity) and high apparent
J
max
.
l
-methionine or
l
-phenylalanine (7 and 20 mM) were competitive inhibitors of
3
H-
l
-leucine influxes in both NaCl and KCl media, producing a significant (
p
< 0.01) increase in
3
H-
l
-leucine influx
K
M
, but no significant response in
3
H-
l
-leucine influx
J
max
. Potassium was a competitive inhibitor of sodium co-transport with
3
H-
l
-leucine, significantly (
p
< 0.01) increasing
3
H-
l
-leucine influx
K
M
in the presence of sodium, but having negligible effect on
3
H-
l
-leucine influx
J
max
in the same medium. These results suggest that shrimp BBMV transport
3
H-
l
-leucine by a single
l
-methionine- and
l
-phenylalanine-shared carrier system that is enhanced by acidic pH and can be stimulated by either Na
+
or K
+
acting as co-transport drivers binding to shared activator sites.
Transepithelial transport of dietary D-glucose and d-fructose was examined in the lobster Homarus americanus intestine using D-(3)Hglucose and D-(3)Hfructose. Lobster intestines were mounted in a ...perfusion chamber to determine transepithelial mucosal to serosal (MS) and serosal to mucosal (SM) transport mechanisms of glucose and fructose. Both MS glucose and fructose transport, as functions of luminal sugar concentration, increased in a hyperbolic manner, suggesting the presence of mucosal transport proteins. Phloridizin inhibited the MS flux of glucose, but not that of fructose, suggesting the presence of a sodium-dependent (SGLT1)-like glucose co-transporter. Immunohistochemical analysis, using a goat anti-rabbit GLUT5 polyclonal antibody, revealed the localization of a brush border GLUT5-like fructose transport protein. MS fructose transport was decreased in the presence of mucosal phloretin in warm spring/summer animals, but the same effect was not observed in cold autumn/winter animals, suggesting a seasonal regulation of sugar transporters. Mucosal phloretin had no effect on MS glucose transport. Both SM glucose and SM fructose transport were decreased in the presence of increasing concentrations of serosal phloretin, providing evidence for the presence of a shared serosal GLUT2 transport protein for the two sugars. The transport of d-glucose and d-fructose across lobster intestine is similar to sugar uptake in mammalian intestine, suggesting evolutionarily conserved absorption processes for these solutes.
The objective of this article is to review the available data on the relationship between sleep-disordered breathing (SDB) and pulmonary arterial hypertension (PAH), with a focus on the prevalence of ...SDB in patients with idiopathic PAH (IPAH); the prevalence of PAH in patients with SDB; and the effects of SDB treatment on PAH. The evidence to date suggests that PAH may occur in the setting of SDB, although the prevalence is low. However, pulmonary hypertension (PH) in SDB is most strongly associated with other risk factors, such as left-sided heart disease, parenchymal lung disease, nocturnal desaturation, and obesity. The limited data available also suggest that SDB is uncommon in patients with IPAH. Treatment of SDB with continuous positive airway pressure may lower pulmonary artery pressures when the degree of PH is mild.
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