Src homology 2-containing protein tyrosine phosphatase-2 acts as a negative regulator for MUC5AC transcription via the inhibition of the ERK1/2 MAPK signalling pathway in the airway.

AIMS: Mucus hypersecretion has been frequently observed in inflammation respiratory diseases. However, the negative regulators for mucus overproduction have not been readily identified. Our work focused on identifying novel negative regulator that modulates mucus overproduction in the human respiratory system. Herein, we examined whether H2 O2 could induce MUC5AC transcription in a dose-dependent manner and activate tyrosine phosphatase (SHP)-2 in human airway epithelial cells. METHODS: We performed qRT-PCR to detect the changes in MUC5AC transcription and dot-blotting analysis to investigate MUC5AC secretion as regulated by SHP-2. RESULTS: H2 O2 induced MUC5AC transcription in a dose-dependent manner and dramatically activated SHP-2. In addition, whereas wild-type SHP-2 completely inhibited H2 O2 -induced MUC5AC transcription, siRNA-SHP-2 restored it interestingly, suggesting that SHP-2 may act as a negative regulator for mucus overproduction and hypersecretion in the human respiratory tract. Moreover, SHP-2 inhibited the ERK1/2 MAPK pathway, thus abolishing the signalling for MUC5AC transcription. CONCLUSION: We found that H2 O2 induced SHP-2 activation, which acted as a suppressor in H2 O2 signalling to regulate MUC5AC transcription in the airway.

c-Ets1 inhibits the interaction of NF-κB and CREB, and downregulates IL-1ß-induced MUC5AC overproduction during airway inflammation.

Mucin hypersecretion is frequently observed in many inflammatory diseases of the human respiratory tract. As mucin hypersecretion refers to uncontrolled mucin expression and secretion during inflammation, studies examining the negative control mechanisms of mucin hypersecretion are vital in developing novel therapeutic medications. We hypothesized that the c-Ets1 induced by interleukin (IL)-1ß would decrease MUC5AC overproduction by inhibiting the interaction of NF-κB with cAMP response element-binding protein (CREB) in vivo. Stimulation with IL-1ß caused the direct binding of NF-κB and CREB to the MUC5AC promoter, thus increasing MUC5AC gene expression. However, IL-1ß-induced MUC5AC messenger RNA levels were surprizingly downregulated by c-Ets1 (located -938 to -930). Interestingly, c-Ets1 also suppressed IL-1ß-induced MUC5AC gene expression in vitro and in vivo by disrupting the interaction of NF-κB with CREB on the MUC5AC promoter. In addition, c-Ets1 also inhibited significant morphologic changes and inflammatory cell infiltration after IL-1ß exposure in mouse lungs infected with either wild-type or shRNA-c-Ets1. Moreover, reactive oxygen species produced by NOX4 increased c-Ets1 gene expression and MUC5AC gene expression in alveolar macrophages from bronchoalveolar lavage fluid. These results suggest a molecular paradigm for the establishment of a novel mechanism underlying the negative regulation of mucin overproduction, thus enhancing our understanding of airway inflammation.

Cadmium inhibits albumin endocytosis in opossum kidney epithelial cells.

Chronic exposure to cadmium results in proteinuria. To gain insights into the mechanism by which cadmium inhibits the protein transport in the renal proximal tubule, we investigated the effects of cadmium on the receptor-mediated endocytosis of albumin, using fluorescein isothiocyanate-labeled bovine serum albumin (FITC-albumin) as a model substrate and opossum kidney cell line (OK cell) as a proximal tubular cell model. Cell monolayers grown to confluence were treated with 100 microM CdCl(2) for 60 min at 37 degrees C, washed, and tested for FITC-albumin uptake (37 degrees C) and surface binding (4 degrees C). The amounts of FITC-albumin uptake and binding were quantified by fluorimetrically determining the cell-adherent fluorescence. Both the binding and uptake of FITC-albumin by OK cells appeared to be saturable and inhibitable by unlabeled albumin in the medium, indicating that specific receptor sites were involved. The uptake of FITC-albumin was inhibited by agents that interfere with the formation of endocytotic vesicle (hypertonic mannitol), endosomal acidification (NH(4)Cl), and vesicular trafficking (cytochalasin D and nocodazole), confirming that the uptake occurred via the process of receptor-mediated endocytosis. In cells treated with cadmium, the specific FITC-albumin uptake was significantly attenuated, and this was due to a reduction in V(max) and a rise in K(m). These changes in kinetic parameters were similar to those induced by NH(4)Cl. The binding of FITC-albumin to the apical surface of OK cells was inhibited by cadmium treatment, and this was attributed to a reduction in B(max). The values of K(d) and its pH dependency were not altered by cadmium treatment. The formation of endocytotic vesicles, as judged by fluid phase endocytosis of FITC-inulin, was not changed by cadmium treatment. These results indicate that the receptor-mediated endocytosis of albumin is impaired in cadmium-treated OK cells most likely due to a defect in endosomal acidification and the attendant fall in ligand-receptor dissociation, which impairs receptor recycling and the overall efficiency of endocytosis.

Cadmium binding and sodium-dependent solute transport in renal brush-border membrane vesicles.

Exposure to cadmium (Cd) impairs renal transport systems for glucose, amino acids, phosphate, and dicarboxylates. To investigate if these changes are directly related to a Cd binding to the renal brush-border membrane, Cd binding and the Na+-dependent uptakes of d-glucose, l-alanine, phosphate, and succinate were determined in rat renal brush-border membrane vesicles (BBMV) exposed to CdCl2. Cd uptake by BBMV showed time and concentration dependence. Changes in medium osmolality had no effect on Cd uptake, indicating that the process primarily involves binding of Cd to the membrane. Scatchard analysis indicated the presence of two types of Cd binding sites, differing in affinity and number. Increasing the medium Cd concentration from 50 to 200 microM resulted in a progressive increase in Cd binding to the membrane and decrease in Na+-dependent transport of d-glucose, l-alanine, inorganic phosphate, and succinate. In all cases, the inhibition of transport was directly proportional to the total amount of Cd binding to the membrane. These results suggest that, during chronic exposure to Cd, free Cd ions liberated in renal tubular cells may directly interact with brush-border membranes and impair Na+-dependent solute transports.

Renal transport systems for organic anions and cations in cadmium-exposed rats.

To evaluate the effect of cadmium intoxication on renal transport systems for organic anions and cations, transport of p-aminohippurate (PAH) and tetraethylammonium (TEA) were studied in renal cortical plasma membrane vesicles isolated from cadmium-intoxicated rats. Cadmium intoxication was induced by daily injections of CdCl2 (2 mg Cd/kg.day sc) for 2-3 weeks. Renal plasma membrane vesicles were prepared by Percoll gradient centrifugation and magnesium precipitation method. Vesicular uptake of substrate was determined by rapid filtration technique using Millipore filter. The cadmium treatment resulted in a marked attenuation of Na(+)-dependent, alpha-ketoglutarate (alpha KG)-driven PAH uptake in the basolateral membrane vesicle (BLMV), and this was due to a reduction in Vmax and not K(m). The Na(+)-alpha KG symport activity of the BLMV was not affected by 2-week cadmium treatment, but it was significantly inhibited by 3-week cadmium treatment. On the other hand, the alpha KG-PAH antiport activity of the BLMV appeared to be markedly suppressed in 2-week as well as 3-week cadmium-treated animals. The cadmium treatment inhibited the proton gradient-dependent TEA transport in the brush-border membrane vesicle (BBMV), and this was associated with a reduction in Vmax with no change in K(m). These results indicate that cadmium exposures may impair the capacities for organic anion transport in the proximal tubular basolateral membrane and organic cation transport in the luminal membrane. The cadmium effect on organic anion transport is attributed mainly to an inhibition of dicarboxylate-organic anion antiport system.

Effect of physical exercise on renal response to head-out water immersion.

Head-out water immersion (HOI) induces various renal functional changes, such as diuresis, natriuresis, and kaliuresis. The present study was undertaken 1) to characterize the renal response to HOI in Koreans who routinely ingest high salt diet and 2) to evaluate the impact of exercise on the renal response to HOI. Six healthy male subjects (average Na+ intake of 232 mEq.day-1) were immersed upto the neck in 34.5 degrees C water and rested in a seated position or exercised on a bicycle ergometer for 3 hours. In resting subjects, we observed a reversible increase in urine flow and a decrease in urine osmolality, with no changes in creatinine clearance. The peak urine flow observed during the second hour of immersion was 4-fold greater than the pre-immersion level. The excretion of total osmotic substances rose progressively during the 3-hour immersion, which was accompanied by a similar change in Na+ excretion. The K+ excretion was slightly elevated. The major component of the immersion diuresis was a water diuresis in the early phase and an osmotic diuresis in the late phase of immersion. In exercising subjects, the diuretic and natriuretic responses to HOI were attenuated and the kaliuretic response was potentiated. Blood hemoglobin concentration and plasma levels of renin, ADH, and aldosterone decreased during immersion-rest, but they remained unchanged or increased during immersion-exercise. These results suggest that 1) the cardiac mechanoreceptor-mediated renal responses to HOI are not changed by chronic high salt diet, and 2) excessive urinary sodium and water losses are prevented by exercise during immersion.

Thermal balance of man in water: prediction of deep body temperature change.

Changes in rectal temperature and metabolic rate were continuously monitored in men immersed in water at 15, 20, 25, 30, and 35 degrees C. The subjects (12 healthy male students) wore swimming suits while either resting or doing leg exercise (delta M approximately 60 kcal.h-1.m-2 approximately 70 W.m-2) for 1 h. At all water temperatures below 30 degrees C, the metabolic rate increased but the rectal temperature fell continuously, resulting in hypothermia. The rate of fall in rectal temperature (cooling rate, CR, degrees C.h-1) was inversely proportional to water temperature (Tw, degrees C) according to the equation CR = 3.818-0.109 Tw in resting subjects and CR = 3.434-0.110 Tw in exercising subjects. At a given Tw the cooling rate was greater in resting than in exercising subjects and in lean than in obese subjects. From the relationships between the cooling rate and water temperature the duration of useful activity and the survival time were predicted for resting and exercising subjects of various fatness.

Transport of inorganic phosphate in renal cortical brush-border membrane vesicles of cadmium-intoxicated rats.

The effect of cadmium intoxication on the renal proximal tubular phosphate transport system was studied in adult male Sprague-Dawley rats. Subcutaneous injections of CdCl2 at a dose of 2 mg Cd/kg body wt per day for 2 weeks induced marked polyuria, glycosuria, proteinuria, and phosphaturia, which are characteristics of chronic cadmium intoxication. In the renal cortical brush-border membrane vesicles prepared from cadmium-intoxicated rats, the cadmium content was drastically increased and the Na(+)-dependent phosphate uptake was markedly attenuated. Similar results were obtained in normal membrane vesicles directly exposed to free cadmium. These results indicate that cadmium intoxication impairs the Na(+)-phosphate cotransport system in the proximal tubular brush-border membrane, which may lead to phosphaturia in intact animals.

Continuous pulse oximetry in the breath-hold diving women of Korea and Japan.

Arterial oxygen saturation during breath-hold diving has not previously been measured continuously. We devised a submersible, waterproof, backpack computer to continuously record heart rate, depth, and arterial oxygen saturation (SPO2) as determined by earlobe pulse oximetry. Our measurements showed that one assisted (Funado) diver had reduced SPO2 values immediately after surfacing from 22 dives which lasted 23-76 s, from a mean of 99 +/- 1% SPO2 to 96 +/- 3% SPO2. SPO2 returned to 97 +/- 2% within 15 s after surfacing (P < 0.05 surface value differs from predive base line). Four unassisted (Cachido) divers showed no significant reduction of mean predive SPO2 below 98 +/- 2% at any time during the dive or recovery period in 92 routine dives lasting from 15 to 44 s. Upon surfacing from diving, mean SPO2 was 98 +/- 2% and the mean SPO2 15 s after surfacing was 97 +/- 3% for the unassisted divers. Three Cachido divers were asked to dive and breath hold for as long as possible. Mean SPO2 at the conclusion of breath holding was 73% after an average dive and breath hold lasting 69 s.

Arterial blood gas tensions during breath-hold diving in the Korean ama.

Korean female unassisted divers (cachido ama) breath-hold dive > 100 times to depths of 3-7 m during a work day. We sought to determine the extent of arterial hypoxemia during normal working dives and reasonable time limits for breath-hold diving by measuring radial artery blood gas tensions and pH in five cachido ama who dove to a fixed depth of 4-5 m and then continued to breath hold for various times after their return to the surface. Eighty-two blood samples were withdrawn from indwelling radial artery catheters during 37 ocean dives. We measured compression hyperoxia [arterial PO2 = 141 +/- 24 (SD) Torr] and hypercapnia (arterial PCO2 = 46.6 +/- 2.4 Torr) at depth. Mean arterial PO2 near the end of breath-hold dives lasting 32-95 s (62 +/- 14 s) was decreased (62.6 +/- 13.5 Torr). Mean arterial PCO2 reached 49.9 +/- 5.4 Torr. Complete return of these values to their baseline did not occur until 15-20 s after breathing was resumed. In dives of usual working duration (< 30 s), blood gas tensions remained within normal ranges. Detailed analysis of hemoglobin components and intrinsic oxygenation properties revealed no evidence for adaptive changes that could increase the tolerance of the ama to hypoxic or hypothermic conditions associated with repetitive diving.

Nitrogen tensions in brachial vein blood of Korean ama divers.

Intravascular bubble formation and symptoms of decompression sickness have been reported during repetitive deep breath-hold diving. Therefore we examined the pattern of blood N2 kinetics during and after repetitive breath-hold diving. To study muscle N2 uptake and release, we measured brachial venous N2 partial pressure (PN2) in nine professional Korean breath-hold divers (ama) during a 3-h diving shift at approximately 4 m seawater depth and up to 4 h after diving. PN2 was determined with the manometric Van Slyke method. Diving time and depth were recorded using a backpack computer-assisted dive longer that allowed calculating the surface-to-depth time ratio to derive the effective depth. With the assumption that forearm muscle N2 kinetics follow the general Haldanian principles of compression and decompression, i.e., forearm muscle is a single compartment with a uniform tissue PN2 equal to venous PN2, PN2 data were fitted to monoexponential functions of time. In the early phase of the diving shift, PN2 rapidly increased to 640 Torr (half time = 6 min) and then slowly declined to baseline levels (half time = 36 min) after the work shift. Peak PN2 levels approximated the alveolar PN2 derived from the effective depth. We conclude that forearm muscle N2 kinetics are well described by a Haldanian single-compartment model. Decompression sickness is theoretically possible in the ama; it did not occur because the absolute PN2 remained low due to the shallow working depth of the ama we studied.

Splenic contraction during breath-hold diving in the Korean ama.

Major increases of hemoglobin concentration and hematocrit, possibly secondary to splenic contraction, have been noted during diving in the Weddell seal. We sought to learn whether this component of the diving response could be present in professional human breath-hold divers. Splenic size was measured ultrasonically before and after repetitive breath-hold dives to approximately 6-m depth in ten Korean ama (diving women) and in three Japanese male divers who did not routinely practice breath-hold diving. Venous hemoglobin concentration and hematocrit were measured in nine of the ama and all Japanese divers. In the ama, splenic length and width were reduced after diving (P = 0.0007 and 0.0005, respectively) and calculated splenic volume decreased 19.5 +/- 8.7% (mean +/- SD, P = 0.0002). Hemoglobin concentration and hematocrit increased 9.5 +/- 5.9% (P = 0.0009) and 10.5 +/- 4% (P = 0.0001), respectively. In Japanese male divers, splenic size and hematocrit were unaffected by repetitive breath-hold diving and hemoglobin concentration increased only slightly over baseline (3.0 +/- 0.6%, P = 0.0198). Splenic contraction and increased hematocrit occur during breath-hold diving in the Korean ama.

Timing of laparoscopic sterilization in abortion patients.

A total of 1604 laparoscopic sterilization procedures with variable time intervals after first trimester therapeutic abortion were performed at the Severance Hospital of the Yonsei University College of Medicine, Seoul, Korea, from May 1973 to October 1975. Two hundred fourteen women were sterilized immediately at the time of abortion; 359 were sterilized between 1 and 42 days later; and the remaining 1031 women were sterilized 43 or more days after abortion. Electrocoagulation and tubal ring application were the tubal occlusion techniques used. The findings indicate that patients who underwent the combined abortion-sterilization procedure did not encounter higher rates of technical problems and/or complications than the other 2 groups. Only a few of these 1604 women studied had potentially serious complications that necessitated subsequent laparotomy, hospitalization after sterilization, and/or hospital readmission.