Volume Reabsorption Research Articles

Socket Preservation Using Dentin Mixed with Xenograft Materials: A Pilot Study.

The use of human dentin matrix could serve as an alternative to autologous, allogenic, and xenogeneic bone grafts due to its osteoinductive characteristics. The limitations of its use is tooth availability and that it is often necessary to mix it with a biomaterial. The aim of this study was to analyze a mix of two different graft materials with different reabsorption ranges when the dentin graft material was not sufficient for full socket preservation. Seven socket preservation surgeries were carried out employing a mixed graft material containing 50% dentin and 50% xenograft. After four months of recovery, the implants were positioned. At the time of the prosthesis placement and implant surgery, bone samples were collected. The histologic analysis revealed no inflammatory or infective reaction against the seven biopsies. The histomorphometric graft analysis revealed an amount of New Bone of 29.03 ± 6.57% after 4 months and 34.11 ± 5.02% after 8 months. The two graft materials had a different volume reabsorption rate: 71% after 4 months and 90% after 8 months for dentin, and 6% after 4 months and 26% after 8 months for the xenograft. The space created by the dentin reabsorption increased the quantity of new bone.

High dietary K+ intake inhibits proximal tubule transport.

The impact of chronic dietary K+ loading on proximal tubule (PT) function was measured using free-flow micropuncture along with measurements of overall kidney function, including urine volume, glomerular filtration rate, and absolute and fractional Na+ and K+ excretion in the rat. Feeding animals a diet with 5% KCl [high K+ (HK)] for 7 days reduced glomerular filtration rate by 29%, increased urine volume by 77%, and increased absolute K+ excretion by 202% compared with rats on a 1% KCl [control K+ (CK)] diet. HK did not change absolute Na+ excretion but significantly increased fraction excretion of Na+ (1.40% vs. 0.64%), indicating that fractional Na+ absorption is reduced by HK. PT reabsorption was assessed using free-flow micropuncture in anesthetized animals. At 80% of the accessible length of the PT, measurements of inulin concentration indicated volume reabsorption of 73% and 54% in CK and HK, respectively. At the same site, fractional PT Na+ reabsorption was 66% in CK animals and 37% in HK animals. Fractional PT K+ reabsorption was 66% in CK and 37% in HK. To assess the role of Na+/H+ exchanger isoform 3 (NHE3) in mediating these changes, we measured NHE3 protein expression in total kidney microsomes as well as surface membranes using Western blots. We found no significant changes in protein in either cell fraction. Expression of the Ser552 phosphorylated form of NHE3 was also similar in CK and HK animals. Reduction in PT transport may facilitate K+ excretion and help balance Na+ excretion by shifting Na+ reabsorption from K+-reabsorbing to K+-secreting nephron segments.NEW & NOTEWORTHY In rats fed a diet rich in K+, proximal tubules reabsorbed less fluid, Na+, and K+ compared with those in animals on a control diet. Glomerular filtration rates also decreased, probably due to glomerulotubular feedback. These reductions may help to maintain balance of the two ions simultaneously by shifting Na+ reabsorption to K+-secreting nephron segments.

3: Vitamin D Improves Autologous Fat Graft Retention

Purpose:Autologous fat grafting is a widely used technique in aesthetic and reconstructive surgery, however, unpredictable volume reabsorption may lead to unsatisfactory outcomes. Previously, we demonstrated that a fat-soluble Vitamin D3 analogue, calcitriol, significantly improved fat retention in a xenograft mouse model by 25% across multiple donors when injected systemically (p < 0.05). While calcitriol has minimal toxicity, is FDA approved, and has positive immunomodulatory and antioxidant properties, systemic administration bypasses key Vitamin D synthesis regulatory steps, thus increasing risk with high-dose use. We hypothesize that systemic supplementation with Vitamin D3 (cholecalciferol) will likewise improve fat-graft retention similar to calcitriol while avoiding potential regulatory and iatrogenic risk. In this study we compared in vivo human fat graft retention in mice treated with systemic cholecalciferol, calcitriol or vehicle control in a mouse xenograft model. In vitro adipose lipoaspirate culture was used to interrogate the therapeutic mechanism of action.Methods:Lipoaspirate was harvested from 6 unique donors using a 2mm cannula and used in parallel for both in vitro and in vivo studies. In vivo: 0.3mL of lipoaspirate was injected bilaterally on dorsal flanks of homozygous Foxn1nu immunocompromised mice. Calcitriol (50ng), cholecalciferol (50ng, 500ng, 5000ng) or vehicle control was administered thrice weekly by IP injection. Graft volume retention was measured at 12 weeks. In vitro: 1mL of lipoaspirate was submerged in phenol-red free DMEM (10% FBS) containing calcitriol or cholecalciferol (15.6nM, 62.5 nM or 250 nM) for 7 days with one media change. Terminal analyses include tissue weight, stromal cell viability, concentration of active vitamin D metabolite (1,25(OH)2D3), and gene upregulation by qRT-PCR.Results:Previously, we demonstrated that systemic administration of 50ng calcitriol thrice weekly significantly improved human fat graft retention across multiple donors in a mouse xenograft model. Our current in vivo data suggest 5000ng cholecalciferol is similarly effective. In-vitro assays show 62.5nM and 250nM cholecalciferol significantly increased adipose stromal cell viability compared to controls (85.3+/-2.9% and 87.7+/-3.7 versus 77.6+/-2.8%, respectively p<0.05;). Analysis of final adipose 1,25(OH)2D3 concentration by ELISA showed both calcitriol and cholecalciferol treatments equally increased Vitamin D metabolite concentrations in all donors. qRT-PCR analysis of gene expression show that pro-survival autophagy is significantly increased by both cholecalciferol and calcitriol, though increased concentration of calcitriol was required to induce significant increases from controls.Conclusion:Cholecalciferol (Vitamin D3) is a highly promising therapeutic for improving fat grafting outcomes. Our in vitro data suggests that in the context of hypoxia, nutrient depletion, or growth factor deprivation, such as occurs immediately following fat grafting, vitamin D3 promotes stromal cell autophagy. In this context, autophagy is crucial for maintaining cellular ATP production and macromolecular synthesis and, therefore, represents an essential pro-survival pathway which allows grafted cells to survive. The results herein provide evidence to incorporate vitamin D3 as a safe, cost-effective nutritional supplement into the perioperative workflow to improve fat viability after grafting.

Functional Implications of Sexual Dimorphism of Transporter Patterns along the Rat Nephron

The goal of this study is to investigate the functional implications of the sexual dimorphism in transporter patterns along the rodent nephron as reported by Veiras et al. (J Am Soc Nephrol 2018). To do so, we developed sex‐specific computational models of water and solute transport from Bowman space to the papillary tip of a superficial nephron of the kidneys of the male and female rats. The models account for the sex differences in expression levels of the apical and basolateral transporters, in single‐nephron glomerular filtration rate, and in tubular dimensions. Model simulations predict that ~70 and 40% of the filtered volume is reabsorbed by the proximal tubule of the male and female rat kidneys, respectively. The lower fractional volume reabsorption in the female rat can be attributed to their smaller transport area and lower aquaporin‐1 expression level. The latter also results in a larger contribution of the paracellular pathway to water transport. Similar fractions of the filtered Na+ are reabsorbed by the male and female proximal tubule models, respectively. The lower fractional Na+ reabsorption in female is due primarily to their smaller transport area, lower Na+/H+‐exchanger (NHE3) activity, and lower claudin‐2 abundance. This results in significantly larger delivery of water and Na+ to the downstream nephron segments in female. Unlike in the proximal tubule, the distal nephron of the female rat exhibits higher expression levels in key Na+ transporters, including Na+‐K+‐Cl− cotransporter, Na+‐Cl− cotransporter, and epithelial Na+ channel. Not only do the higher expression levels of these transporters account for the enhanced water and Na+ transport along the female distal nephron, relative to male, but they may also contribute to the female rat's ability to excrete a saline load more rapidly than male.Support or Funding InformationThis research was supported by the Canada 150 Research Chair program and by the National Institutes of Health: National Institute of Diabetes and Digestive and Kidney Diseases, grant R01DK106102.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The Use of Double-Layer Collagen Membrane for the Improvement of Nasal Dorsum Skin Thickness and Texture in Primary Nose Surgery.

To place a collagen membrane containing crushed nasal septal cartilage over the nasal dorsum to see how this graft can improve the results of visible postsurgical irregularities in thin skinned patients. Fifty-seven patients were treated between 2006 and 2010 (26 males and 31 females) whose ages ranged between 31 and 55 years old. They were divided into group a, defect <1 mm, group b defect between 1 and 2 mm, group c defect >3 mm. These patients presented intraoperative nasal dorsum irregularities that were corrected with 57 cartilage grafts in conjunction with collagen membrane. All of our patients showed an initial over-correction. A dense network of collagen fiber bundles was observed running parallel to the surface of the membrane at the connective tissue-membrane interface. The clinical percentage of volume reabsorption was about 5% after 6 months, measured with clinical pictures and a clinical follow-up. No infection was noted, only 1 patient of dislocation was observed, and 5 patients required some refinements at the long-term follow-up. In the authors' experience the use of bilayered combined cartilage and collagen membrane grafts gives the best aesthetic results with balanced tip projection and dorsum fullness, and avoids thickness and texture modification of the skin above the graft. The use of Bio-gide membrane avoids all problems related to the donor site and shortens surgical time.

Sexual Dimorphic Expression of Renal Claudins, Water Channels and Transporters accounts for the Downstream Shift in Salt and Volume Reabsorption along the Nephron in Female vs. Male Rats

Recently, we reported that female rats, as well as mice, have lower abundance of apical proximal tubule (PT) sodium transporters (NHE3, NaPi2) as well as lower abundance of myosin VI, the motor that drives regulated redistribution of the apical transporters. This reduction in PT transporters in females is complemented by higher abundance and phosphorylation of distal tubule Na‐Cl cotransporter (NCC) and higher abundance of cleaved (activated) forms of epithelial sodium channels’ (ENaC) alpha and gamma subunits. At the physiologic level, the impact of the differences is evident as more rapid excretion of a saline load, higher clearance rate of endogenous lithium (a marker of flow from the proximal tubule), and no difference in thiazide sensitive natriuresis in females vs. males. The aim of the current study was to test the hypothesis that a sexual dimorphism in claudins (Cld‐) and water channels (AQP‐) accompanies the observed differences in transporters. Sprague Dawley rats were fasted overnight with water provided before termination. Abundance of claudins and water channels were determine by quantitative immunoblot of cortical or medullary homogenates. Claudin‐2, expressed from the PT through the thin descending limb, forms high‐conductance cation‐selective paracellular pores that play an important role in the paracellular reabsorption of salt in the PT (&gt;32% of the filtered load). Claudin‐2 expression in cortex was lower in females vs. males: 0.32 ± 0.03 vs. 1.0 ± 0.1 (p&lt; 0.002). Aquaporin 1, which confers “leakiness” to the PT, expressed in both apical and basolateral membranes of the PT, was also lower in females vs. males: 37 kD band: 0.44 ± 0.014 vs. 1.0 ± 0.023; 25 kD band: 0.84 ± 0.22 vs. 1.0 ± 0.26 (both p&lt;0.001). Claudin‐7, expressed in the DCT and CCD, functions as either a Cl− barrier or pore; in vivo knockout provokes lethal salt wasting. Claudin‐7 expression in cortex was higher in females vs. males: 1.4 ± 0.01 vs. 1.0 ± 0.03 (p&lt; 0.001). Claudin‐10, detected all along the nephron, and medullary apical Aquaporin‐2 were expressed at similar levels in females and males.ConclusionsLower abundance of both Cld‐2 and AQP‐1 in females corresponds to lower Na+ transporter abundance in the proximal nephron and may contribute to their improved ability to excrete a saline load. These factors may contribute to the lower blood pressures and resistance to experimental hypertension in females vs. males. In the distal nephron, higher abundance of Cld‐7 corresponds to the higher NCC and ENaC activities where it may facilitate salt reabsorption in response to the increased volume flow from the proximal nephron.Support or Funding InformationAHA 15GRNT23160003 and NIH DK083785

REGULATION OF GLOMERULOTUBULAR BALANCE IV: IMPLICATION OF AQUAPORIN 1 IN FLOW‐DEPENDENT TRANSPORT AND CELL VOLUME

Underlying glomerulortubular balance (GTB) is the impact of axial flow to regulate Na+ and HCO3− transport by modulating luminal NHE3 and H‐ATPase, and presumably other peritubular transporters, since there is little flow‐dependent change in cell volume. Aquaporin (AQP‐1) serves as a water channel and is the principal water pathway for luminal and peritubular membranes of proximal tubule. We investigated flow‐mediated fluid (Jv) and HCO3− (JHCO3) absorption in proximal tubules (S2) of mouse kidney by microperfusion in vitro in wild‐type (WT) and AQP‐1 KO mice. Perfusion Tubule No. Jv (nl/min/mm) JHCO3 (pmol/min/mm) Cell Volume (μm3) (nl/min) WT KO WT KO WT KO WT KO 5 31 9 0.79±0.04 0.57±0.06† 56.3±3.1 61±5.9 1027 1034 20 10 18 1.37±0.06* 0.38±0.03*† 116±7.5* 133±7.8* 1001 946 P&lt;0.05 compared from low flow rate; P&lt;0.05 compared between WT and KO at same flow rate. An increase in perfusion rate from 5 to 20 nl/min increased Jv by 73% and JHCO3 by 106% in proximal tubules of WT mice (P&lt;0.001). Compared with WT, AQP‐1 KO significantly decreased Jv at both low and high flow rates (P&lt;0.05), and there was actually a decrease in volume flux in the transition from low to high flow (P&lt;0.05). In contrast, the AQP‐1 KO mice showed no difference from WT in JHCO3 at either low or high flow rates. Cell volume showed no significant difference between either low or high flow rates or between WT and AQP‐1 KO mice. These results confirm the role of AQP‐1 as the principal pathway for water reabsorption in proximal tubule. The decrease in Jv with increasing perfusion rate is consistent with a model in which proximal Na+ reabsorption produces a transepithelial osmotic gradient, which then drives volume reabsorption. Whether axial flow increases AQP‐1 activity, remains uncertain. Flow‐stimulated Na+ and HCO3− absorption is independent of intact AQP‐1.

Proximal tubule specific knockout of the Na+/H+ exchanger NHE3: effects on bicarbonate absorption and ammonium excretion

The existing NHE3 knockout mouse has significant intestinal electrolyte absorption defects, making this model unsuitable for the examination of the role of proximal tubule NHE3 in pathophysiologic states in vivo. To overcome this problem, we generated proximal convoluted tubule-specific KO mice (NHE3-PT KO) by generating and crossing NHE3 floxed mice with the sodium-glucose transporter 2 Cre transgenic mice. The NHE3-PT KO mice have >80 % ablation of NHE3 as determined by immunofluorescence microscopy, western blot, and northern analyses, and show mild metabolic acidosis (serum bicarbonate of 21.2 mEq/l in KO vs. 23.7 mEq/l in WT, p < 0.05). In vitro microperfusion studies in the isolated proximal convoluted tubules demonstrated a ∼36 % reduction in bicarbonate reabsorption (J HCO3 = 53.52 ± 4.61 pmol/min/mm in KO vs. 83.09 ± 9.73 in WT) and a ∼27 % reduction in volume reabsorption (J v = 0.67 ± 0.07 nl/min/mm in KO vs. 0.92 ± 0.06 nl/min/mm in WT) in mutant mice. The NHE3-PT KO mice tolerated NH4Cl acid load well (added to the drinking water) and showed NH4 excretion rates comparable to WT mice at 2 and 5 days after NH4Cl loading without disproportionate metabolic acidosis after 5 days of acid load. Our results suggest that the Na+/H+ exchanger NHE3 plays an important role in fluid and bicarbonate reabsorption in the proximal convoluted tubule but does not play an important role in NH4 excretion.

Effect of sulodexide on peritoneal morphology and function of peritoneal dialysis rats

Objective To investigate the effect of sulodexide on the peritoneal morphology and function of long-term peritoneal dialysis rats.Methods Adult male Sprague-Dawley rats were randomly divided into the following four groups: control group(n=6),model group(n=10),low-dose sulodexide group(n=10) and high-dose sulodexide group(n=10).A 1 hour peritoneal equilibration test was performed after an eight-week peritoneal dialysis in each group.The dialysate samples were subjected to examination by hemocytometer for determination of 1 h dialysate urea(Durea),dialysate protein(Dtp),initial dialysate glucose(D0) and 1 h dialysate glucose(D1),plasma urea nitrogen(Purea)and total plasma protein(Ptp);and the D/Purea(parameter for clearance rate of urea nitrogen),D/Ptp(parameter for total protein loss in dialysate),and D1/D0(parameter for peritoneal ultrafiltration)were calculated.The parietal peritoneum tissues of rats were harvested and stained by Hematoxylin-eosin and Masson trichrome for observation of the peritoneal structure.The vessels and leukocytes in peritoneum tissue were counted(n/mm2) using stained sections.Leukocytes in dialysate samples were evaluated using hemocytometer.The levels of inflammatory cytokine(MCP-1,TNFα) in dialysate samples were measured by enzyme-linked immunosorbent assay(ELISA) and the expression of TGF-β1 in parietal peritoneum was detected by immunohistochemistry assay.Results Compared with the control group,rats in the model group had decreased peritoneal mesothelial cells,thickened sub-mesothelial matrix,increased inflammatory cells,and angiogenesis(P0.05).The ultrafilitration volume(UF) and glucose reabsorption(D1/D0) were significantly lower and the dialysate-to-plasma urea ratio(D/Purea) and dialysate-to-plasma total protein ratio(D/Ptp) were significantly higher in model group than those in the control group(P0.05).Counts of leukocytes and levels of MCP-1 and TNFα in dialysate samples were significantly increased(P0.05) and TGF-β1 expression in parietal peritoneum was also significantly increased in the model group.Administration of sulodexide greatly improved the structure changes of the peritoneum,decreased the inflammatory cell infiltration and angiogenesis,greatly improved peritoneal function,significantly decreased the levels of MCP-1 and TNF-α in dialysate samples(P0.05),and decreased the expression of TGF-β1 in parietal peritoneum.Conclusion Sulodexide may improve the peritoneal morphology and function by inhibiting pertioneal chronic inflammation.

Glomerular filtration rate in early diabetes: ongoing discussions of causes and mechanisms

Glomerular hyperfiltration (>140 ml/min per 1.73 m2 body surface area) is found in early diabetes and is associated with a poor prognosis with respect to the development of diabetic kidney disease. This review addresses recent investigations and discussions of the following hypotheses behind diabetic hyperfiltration: Increased proximal tubular volume reabsorption results in a pressure drop in Bowman's capsule which increases glomerular filtration rate (GFR). Proximal tubular hyperreabsorption induces an increase in GFR mediated by tubuloglomerular feedback. Dietary NaCl restriction results in a paradoxically increased GFR and increased urine volume in diabetic animals.

Shear stress-induced changes of membrane transporter localization and expression in mouse proximal tubule cells

Our previous studies of microperfused single proximal tubule showed that flow-dependent Na(+) and HCO(3)(-) reabsorption is due to a modulation of both NHE3 and vacuolar H(+)-ATPase (V-ATPase) activity. An intact actin cytoskeleton was indicated to provide a structural framework for proximal tubule cells to transmit mechanical forces and subsequently modulate cellular functions. In this study, we have used mouse proximal tubule (MPT) cells as a model to study the role of fluid shear stress (FSS) on apical NHE3 and V-ATPase and basolateral Na/K-ATPase trafficking and expression. Our hypothesis is that FSS stimulates both apical and basolateral transporter expression and trafficking, which subsequently mediates salt and volume reabsorption. We exposed MPT cells to 0.2 dynes/cm(2) FSS for 3 h and performed confocal microscopy and Western blot analysis to compare the localization and expression of both apical and basolateral transporters in control cells and cells subjected to FSS. Our findings show that FSS leads to an increment in the amount of protein expression, and a translocation of apical NHE3 and V-ATPase from the intracellular compartment to the apical plasma membrane and Na/K-ATPase to the basolateral membrane. Disrupting actin by cytochalasin D blocks the FSS-induced changes in NHE3 and Na/K-ATPase, but not V-ATPase. In contrast, FSS-induced V-ATPase redistribution and expression are largely inhibited by colchicine, an agent that blocks microtubule polymerization. Our findings suggest that the actin cytoskeleton plays an important role in FSS-induced NHE3 and Na/K-ATPase trafficking, and an intact microtubule network is critical in FSS-induced modulation of V-ATPase in proximal tubule cells.

Renal Fanconi Syndrome Secondary to Deferasirox: Where There Is Smoke There Is Fire

The kidney filters ~150 liters per day and reabsorbs 99% of this ultrafiltrate of plasma. The renal tubule is segmented and different portions of the nephron have selective roles in solute and volume reabsorption. The proximal tubule reabsorbs four-fifths of the filtered bicarbonate, two-thirds of the filtered chloride, and essentially all of the filtered glucose, amino acids, and phosphate. Thus, significant injury to the proximal tubule leads to volume depletion, metabolic acidosis, and hypophosphatemia. Hypokalemia is also a common manifestation as aldosterone levels are elevated due to volume depletion and there is increased delivery of sodium and bicarbonate to the distal nephron where sodium is reabsorbed causing a lumen negative potential leading to potassium secretion across an apical potassium channel. Hypomagnesemia is also a common feature of the Fanconi Syndrome.

Osteogenesis distraction and platelet‐rich plasma: combined use in restoration of severe atrophic mandible. Long‐term results

The purpose of this paper is to report long-term results on the use of autologous bone graft and platelet-rich plasma in alveolar distraction osteogenesis (DO) for restoration of severe atrophic mandible. We tested the efficacy as to reabsorption of bone volume, peri-implant reabsorption, implant survival and success rate. Twelve patients were treated. The surgical procedure consisted in mixing autologous bone, harvested from the iliac crest, with autologous platelet concentrate (APC) and in filling the distraction gap with this graft. After a latency of 15 days, a distraction rate of 0.5 mm/day was followed. After a 60-day period of consolidation, the distraction device was removed and implants were placed simultaneously. The abutment connection was accomplished after 6 months. In addition, every patient was evaluated clinically and radiographically annually for 5 years. Planned alveolar height was reached in 11 out of 12 patients. The total number of implants positioned was 47. At the time of implant positioning, the mean decrease of total bone volume was 2.3%. The mean peri-implant resorption was 0.40 mm at the time of abutment connection, 0.61 mm 1 year after implant loading and 1.51 mm after 5 years. After 5 years of follow-up, the mean rate of vertical bone loss was 18.7%. Instead, the implant survival and success rates were 97.9% and 91.5%, respectively. Long-term results allow us to confirm the combination of autologous bone-platelet gel with alveolar DO as an effective and predictable procedure in restoration of severe atrophic mandible.

INTERNALIZATION OF DCT Na‐Cl‐CO‐TRANSPORTER (NCC) DURING PRESSURE DIURESIS‐NATRIURESIS

When blood pressure (BP) is elevated above baseline, a pressure natriuresis response ensues which is critical to volume and blood pressure homeostasis. The response involves a decrease in salt and volume reabsorption from proximal tubule and a load dependent increase in salt reabsorption in loop of Henle. The role of distal nephron is less clear. Distal convoluted tubule Na+‐Cl− cotransporter (NCC) is regulated by trafficking between apical plasma membrane (APM) and sub‐apical cytoplasmic vesicles (SCV). We aimed to determine whether NCC trafficking contributes to pressure natriuresis (increase APM NCC) or compensates for increased volume flow to the DCT (decrease APM NCC). BP was raised 50 mmHg (hiBP) for 30 min in rats by arterial constriction. Kidneys were excised and NCC subcellular distribution analyzed on sorbitol density gradients by immunoblot or kidneys were perfusion fixed and NCC analyzed by immuno‐electron microscopy. After 30 min hiBP urine output increased 11‐fold. By density gradient, 20% of the total NCC redistributed from low density APM to higher density fractions (p&lt;0.05). By quantitative immuno‐EM pool size of APM NCC decreased 13% with a corresponding increase in the SCV pool (p&lt;0.05). In summary, the results provide evidence for an acute decrease in APM NCC during hypertension that likely contributes to pressure diuresis‐natriuresis. HL085388, DK34316, Danish NRF.

Fluid transport and ion fluxes in mammalian kidney proximal tubule: a model analysis of isotonic transport

By mathematical modelling, we analyse conditions for near-isotonic and isotonic transport by mammalian kidney proximal tubule. The model comprises compliant lateral intercellular space (lis) and cells, and infinitely large luminal and peritubular compartments with diffusible species: Na+, K+, Cl- and an intracellular non-diffusible anion. Unknown model variables are solute concentrations, electrical potentials, volumes and hydrostatic pressures in cell and lis, and transepithelial potential. We used data mainly from rat proximal tubule to model epithelial cells and interspace with luminal and peritubular baths of identical composition. The model of the tubular epithelium with physiological water permeability and paracellular electrical resistance generates solute coupled water uptake with an approx. 3% hypertonic absorbate. This function remains unperturbed following 'blocking' of apical water channels and in 'aquaporin-null' simulation. Reduced rate of volume reabsorption in AQP(-/-) mice would also require decreased apical sodium permeability. Paracellular convection accounts for approx. 36% of the net Na+ absorption, and the model epithelium accomplishes uphill water transport similar to rat proximal tubule. Na+ recirculation is required for truly isotonic transport. The tonicity of the absorbate and the recirculation flux depend critically on ion permeabilities of interspace basement membrane. Our model based on solute-solvent coupling in lateral space simulates major physiological features of proximal tubule, including significantly lower water permeability of the AQP1-null preparation, and a ratio of net sodium uptake and oxygen consumption exceeding that predicted from stoichiometry of the Na+/K+-pump. Physical properties of interspace basement membrane are critical for obtaining near-isotonic and truly isotonic transport.

ACE inhibitors’ use in preventing and managing heart disease

Clinical trials have shown that angiotensin converting enzyme (ACE) inhibitors play an important role in the management of patients with, or at risk of heart disease, in particluar those with hypertension, heart failure and ventricular dysfunction after a myocardial infarction. The following paper will review the relevant physiology and pharmacology of ACE inhibitors and present the research evidence of this drug group in relation to these patients. It can be seen that the benefit of ACE inhibitors lies in their ability to reduce vasoconstriction and volume reabsorption through the blockade of Angiotensin II production and the positive effects this has on bradykinin activities.

Proximal tubule water transport-lessons from aquaporin knockout mice

THE PROXIMAL TUBULE REABSORBS essentially all the filtered organic solutes, most of the filtered phosphate, 80% of the filtered bicarbonate, and 60% of the filtered sodium chloride. Approximately 70% of the filtered water is also reabsorbed by this segment. Despite these very high rates of proximal tubule solute transport, the osmolality of the luminal fluid decreases by only 5 mosmol/kgH2O from Bowman’s space to the end of the proximal tubule accessible by micropuncture (3). Furthermore, the luminal osmolality was found to be 7.5 mosmol/ kgH2O lower than that in the peritubular plasma in MunichWistar rats (3). This luminal hypotonicity provides the driving force for water reabsorption in the proximal tubule. The fact that there is only a very small decrease in luminal osmolality is consistent with a very high diffusional water permeability in this nephron segment (10). Despite much debate, physiological studies showed that the vast majority of water is transported across the proximal tubular cell and not across the paracellular pathway (1, 8, 10). The mechanism for this high rate of transcellular water flow was elusive until the cloning of aquaporin-1 (9), which was found to be expressed in high abundance on the apical and basolateral membranes of the proximal tubule and also in the thin descending limb (7). Since these seminal observations, a family of aquaporins has been cloned. At least seven aquaporin isoforms are expressed in the kidney, and each different nephron segment has a unique aquaporin isoform expression (6). In addition to aquaporin-1, aquaporin-7 is also expressed on the proximal tubule but only on the apical membrane of the distal portion of the proximal straight tubule, the S3 segment. The role of aquaporin-1 in mediating proximal tubule water transport has been delineated by elegant studies by Schnermann, Verkman, and co-workers (11). Aquaporin-1 null mice have an 50% lower rate of proximal tubule volume absorption measured in vivo and in vitro than do wild-type mice (11). These data indicate that water permeability can be a limiting factor in proximal tubule solute transport. These mice also have a lower brush-border membrane vesicle osmotic water permeability compared with wild-type mice (4). In addition, aquaporin-1 null mice had an 80% reduction in transepithelial water permeability consistent with most of water movement occurring transcellularly via aquaporin-1 (11). Aquaporin-1 null mice generate a higher proximal tubule transepithelial osmotic gradient despite the lower rates of volume reabsorption, indicating that the nearly isotonic reabsorption of proximal tubular fluid is largely due to water movement through aquaporin-1 (13). Aquaporin-1 null mice have polydipsia and polyuria and an impaired concentrating ability despite the fact that the collecting duct expresses the vasopressin-responsive aquaporin-2 on the apical membrane and aquaporins-3 and -4 are present on the basolateral membrane (5, 11). The impaired concentrating mechanism in aquaporin-1 null mice is probably not secondary to an increased solute load from impaired proximal tubule transport flooding the distal nephron as tubular glomerular feedback decreases single-nephron GFR in these mice (11) but is most likely the result of an inability to maximally form a hypertonic medulla due to the lack of aquaporin-1 in the thin descending limb. The study by Sohara et al. (12) examined the importance of aquaporin-7 in water and glycerol transport using aquaporin-7 knockout mice. Aquaporin-7, unlike aquaporin-1, also facilitates glycerol as well as water transport. Brush-border membrane vesicles from the outer medulla, which includes the S3 proximal tubule, of aquaporin-7 knockout mice had only a 10% reduction in osmotic water permeability. Thus even in the late proximal straight tubule, aquaporin-1 appeared to be the channel responsible for most water transport. Furthermore, unlike aquaporin-1 knockout mice (4), aquaporin-7 null mice did not have a reduction in urinary concentrating ability. Rather than assume that aquaporin-7 had no significant role in proximal tubule water reabsorption, the authors generated an aquaporin1/aquaporin-7 double-knockout mouse. They found that brush-border membrane vesicles from the outer medulla of double-knockout mice had a lower osmotic permeability than that of aquaporin-1 knockout mice. Furthermore, the doubleknockout mouse had a greater impairment in urinary concentrating ability compared with the aquaporin-1 knockout mouse, indicating that the normal urinary concentrating ability in the aquaporin-7 knockout was due to compensation by aquaporin-1 and that aquaporin-7 does indeed play an important role in proximal straight tubule water reabsorption.

Evidence from renal proximal tubules that \documentclass[12pt]{minimal}\usepackage{amsmath}\usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy}\usepackage{mathrsfs}\setlength{\oddsidemargin}{-69pt}\begin{document}\begin{equation*}{\mathrm{HCO}}_{3}^{-}\end{equation*}\end{document} and solute reabsorption are acutely regulated not by pH but by basolateral \documentclass[12pt]{minimal}\usepackage{amsmath}\usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy}\usepackage{mathrsfs}\setlength{\oddsidemargin}{-69pt}\begin{document}\begin{equation*}{\mathrm{HCO}}_{3}^{-}\end{equation*}\end{document} and CO2

Respiratory acidosis, a decrease in blood pH caused by a rise in [CO(2)], rapidly triggers a compensatory response in which the kidney markedly increases its secretion of H(+) from blood to urine. However, in this and other acid-base disturbances, the equilibrium CO(2) + H(2)O HCO(3)(-) + H(+) makes it impossible to determine whether the critical parameter is [CO(2)], [HCO(3)(-)], and/or pH. Here, we used out-of-equilibrium CO(2)/HCO(3)(-) solutions to alter basolateral (BL) [HCO(3)(-)], [CO(2)], or pH, systematically and one at a time, on isolated perfused S2 rabbit proximal tubules. We found that increasing [HCO(3)(-)](BL) from 0 to 44 mM, at a fixed [CO(2)](BL) of 5% and a fixed pH(BL) of 7.40, caused HCO(3)(-) reabsorption (J(HCO(3))) to fall by half but did not significantly affect volume reabsorption (J(V)). Increasing [CO(2)](BL) from 0% to 20%, at a fixed [HCO(3)(-)](BL) of 22 mM and pH(BL) of 7.40, caused J(HCO(3)) to rise 2.5-fold but did not significantly affect J(V). Finally, increasing pH(BL) from 6.80 to 8.00, at a fixed [HCO(3)(-)](BL) of 22 mM and [CO(2)](BL) of 5%, did not affect either J(HCO(3)) or J(V). Analysis of the J(HCO(3)) and J(V) data implies that, as the tubule alters J(HCO(3)), it compensates the reabsorption of other solutes to keep J(V) approximately constant. Because the cells cannot respond acutely to pH changes, we propose that the responses of J(HCO(3)) and the reabsorption of other solutes to changes in [HCO(3)(-)](BL) or [CO(2)](BL) involve sensors for basolateral HCO(3)(-) and CO(2).

Renal water reabsorption: A physiologic retrospective in a molecular era

The cloning and sequencing of the aquaporin water channels has been an enormous advance in the biomedical sciences, as recognized by the award of the Nobel Prize to Peter Agre last year. Among many other examples, expression of aquaporin proteins in Xenopus oocytes and other heterologous expression systems has confirmed two important models of renal function: the increase in the water permeability of the collecting duct by antidiuretic hormone (ADH), and the mechanism of near isosmotic volume reabsorption by the proximal tubule. These mechanisms were the subjects of intensive investigation by numerous investigators, including Thomas E. Andreoli, who is being honored by this symposium, and who developed many of the key concepts in these areas. His early work with artificial lipid bilayer membranes and the pore-forming antibiotic amphotericin provided the rigorous foundation in experimental and conceptual modeling techniques that he later applied to physiologic and pathophysiologic mechanisms in the kidney, which are summarized in this retrospective. Dr. Andreoli and his colleagues proposed a water channel mechanism for the action of ADH, which has been confirmed by the cloning and heterologous expression of aquaporin-2. They also proposed that volume reabsorption by the proximal tubule depended on a very high hydraulic conductivity and the development of luminal hypotonicity produced by active solute reabsorption. This model has also been confirmed in mice in which aquaporin-1 expression is knocked out, resulting in a low proximal tubule water permeability that exaggerates the development of luminal hypotonicity.

Mendelian hypertension with brachydactyly as a molecular genetic lesson in regulatory physiology.

Mendelian forms of hypertension have delivered a treasure trove of novel genes. To date, the molecular mechanisms of five such syndromes have been largely clarified, including glucocorticoid-remediable aldosteronism, Liddle's syndrome, apparent mineralocorticoid excess, an activating mutation of the mineralocorticoid receptor, and pseudohypoaldosteronism type 2. Each of these conditions features salt sensitivity with increased sodium and volume reabsorption by the kidney and low plasma renin activity. None of the gene loci for these syndromes has been convincingly linked to hypertension in the general population. We are investigating kindreds who have autosomal-dominant hypertension and brachydactyly. Affected persons invariably have both anomalies. The hypertension is severe and results in death at about age 50 years from stroke. The condition resembles essential hypertension, because renin, aldosterone, and norepinephrine responses are normal and no salt sensitivity is present. The response to antihypertensive drugs is general. Another feature is diminished baroreflex sensitivity with markedly impaired blood pressure buffering. Furthermore, the ventrolateral medulla may be compromised in these patients, because neurovascular anomalies are a regular finding. We mapped the gene(s) for this disease to chromosome 12p and narrowed the chromosomal region by studying more affected families. Interestingly, the same locus was recently mapped in Chinese families with essential hypertension. Our 3-centimorgan region contains genes encoding a phosphodiesterase, an ATP-dependent potassium channel, and its regulator the sulfonylurea receptor 2. Screening of the coding regions revealed that none of these candidate genes harbor obvious mutations; however, other genetic mechanisms may nevertheless compromise their function. Our study underscores the importance of regulatory physiology to the understanding of a complex genetic syndrome.