CHARACTERISTICS OF MEMBRANE TRANSPORT PROCESSES OF MACULA DENSA CELLS

1. Macula densa (MD) cells are located within the thick ascending limb (TAL) and have their apical surface in contact with tubular fluid and their basilar region in contact with the glomerulus. These cells sense changes in luminal fluid sodium chloride concentration ([NaCl]) and transmit signals resulting in changes in vascular resistance (tubuloglomerular feedback) and renin release. 2. Current efforts have focused on understanding the cellular transport mechanisms of MD cells. Progress in this area has benefited from the use of the isolated perfused TAL-glomerular preparation, which permits direct access to MD cells. 3. Using microelectrodes to measure basolateral membrane potential (V_BL) of MD cells, it was found that VBL was very sensitive to changes in luminal fluid [NaCl]. As [NaCl] was elevated from 20 to 150mmol/L, V_BL was found to depolarize by over 30 mV. 4. Basolateral membrane potential measurements were also used to identify an apical Na⁺: 2CI^?: K⁺ cotransport pathway in MD cells that is the major pathway for NaCl entry into these cells. 5. Other work identified a basolateral chloride channel that is presumed to be responsible for changes in V_BL during alterations in luminal [NaCl]. This channel, which is the predominant conductance across the basolateral membrane, may be regulated by intracellular Ca²⁺ and cAMP. 6. An apical Na⁺: H⁺ exchanger in MD cells was detected by measuring changes in intracellular pH using the fluorescent probe 2′,7′-bis-(2-carboxyethyl)-5(and-6) carboxyfluorescein. 7. Using patch-clamp techniques, a high density of pH- and Ca²⁺-sensitive K⁺ channels was observed at the apical membrane of MD cells. 8. Other studies found that, at the normal physiological conditions prevailing at the end of the TAL (luminal [NaCl] of 20–60 mmol/L), reabsorption mediated by MD cells is very sensitive to changes in luminal [NaCl].     

HFE codon 63/282 (H63D/C282Y) dimorphism in German patients with genetic hemochromatosis

Abstract: Genetic hemochromatosis (GH) is closely associated with genes of the major histocompatibility complex (MHC) on chromosome 6. Recently, a candidate gene for GH, with structural similarities to MHC class I genes, designated HLA-H and presently named HFE, has been cloned. The HFE gene is localized telomeric to the MHC and several reports have indicated that the HFE gene is mutated in GH patients. In the present study we have analyzed the relationship of HFE gene variants and disease manifestation in GH patients and family members. Fifty-seven patients with GH, 73 family members and 153 healthy blood donors were studied for the amino acid dimorphism at codon 63 (His63Asp=H63D) and codon 282 (Cys282Tyr= C282Y) of the HFE gene. The codon 63 and 282 dimorphism were defined by PCR amplification of genomic DNA samples and restriction enzyme digestion using RsaI/SnaBI for C282Y and Bcll/Mbo 1 for H63D. Ferritin, transferrin serum levels and total iron-binding capacity were determined prior to therapeutic intervention. The Tyr-282 substitution occurred in 53 (93%) of patients compared with 8 (5.2%) of controls (OR=169, P >0.0001). Fifty-one (90%) patients were Tyr-282 homozygous. In contrast, the Asp-63 substitution was present in 5 (8.8%) of the patients compared with 34 (22%) of controls (OR=0.39, P =NS) with none of the patients being homozygous. In Tyr-282 homozygous GH patients serum ferritin levels, transferrin saturation, liver iron and liver iron index were elevated significantly compared to Tyr-282-negative patients, whereas no difference was observed between Tyr/Cys-282 heterozygous and Tyr-282-negative patients.     

EFFECT OF SOME TRICYCLIC AND NONTRICYCLIC ANTIDEPRESSANTS ON [3H]IMIPRAMINE BINDING AND SEROTONIN UPTAKE IN RAT CEREBRAL CORTEX AFTER PROLONGED TREATMENT

Chronic administration of different antidepressant drugs reduced the number of [3H]imipramine [( 3H]IMI) binding sites in rat cerebral cortex. In the same experimental conditions, fluvoxamine and dothiepin, as well as desmethylimipramine, induced an increase in the maximal velocity of high affinity serotonin (5HT) uptake in cortical slices, whereas citalopram and viloxazine were ineffective in this regard. Our results indicate that even if 5HT uptake and [3H]IMI binding sites are located on the same nerve terminals, they are differently modulated. Increased Vmax of the 5HT uptake process could be due to a rebound phenomenon after withdrawal from drugs that acutely inhibit 5HT uptake. The effect on [3H]IMI sites might be explained through either the agonist properties of the drugs towards these sites or the involvement of mechanisms still unknown.     

Cytoprotective effect of epidermal growth factor on acid- and pepsininduced damage to rat gastric epithelial cells: Roles of Na+/H+ exchangers

We have previously established a cell damage model, with damage induced by either acid or pepsin treatment for 30 min, involving a rat gastric epithelial cell line (RGM1). In the present study, pretreatment of cells with epidermal growth factor (EGF; 0.1–10ng/mL) or sucralfate (0.1–3 mg/mL) for 4 h prevented such cell damage in a concentration-dependent manner. Protection of cells by these drugs was not affected by pretreatment with indomethacin (10⁻⁵ mol/L) for 4 h. Removal of Na⁻, but not Ca²⁺, from the acidified medium totally abolished the inhibitory effect of EGF, but not that of sucralfate. Genistein (a tyrosine kinase inhibitor) apparently reduced the inhibitory effect of EGF. DNA synthesis by RGM1 cells did not increase when cells were incubated with EGF for 4 h. We conclude that both EGF and sucralfate protect RGM1 cells from acid- and pepsin-induced damage and that the mechanism of protection by EGF against acid-induced damage seems to be via activation of Na⁺/H⁺ exchangers.     

Influence of peripheral afferents on cortical and spinal motoneuron excitability.

The objective of this study was to establish to what extent muscle, cutaneous, and joint afferents alter the excitability of spinal and cortical motor neurons. This question was examined by studying the impact of electrical stimulation of the second and third digits, the median nerve at the wrist, and the recurrent thenar motor branch on the F/H and magneto-electrical cortical motor responses (MEPs) of the thenar muscles. The firing frequencies of single F/H motor unit action potentials were unaltered by the foregoing conditioning peripheral stimuli. MEPs conditioned by motor threshold stimulation of the median nerve at the wrist or the recurrent motor branch were significantly increased in size at conditioning to test intervals of 50 to 80 milliseconds. No significant change in MEP size resulted from conditioning stimulation of the digital nerves. We conclude that muscle afferents were primarily responsible for the increase in MEP size. Conditioning stimuli may allow examiners to assess central motor conduction where it would otherwise be impossible.     

磺胺噻二嗪硫酮衍生物的合成及其抑菌活性

利用药物化学骈合原理设计并合成了一系列新的３，５－二取代１，３，５－噻二嗪－２－硫酮类化合物，其结构经红外光谱，紫外光谱及元素分析证实，抑菌活性试验显示了良好的抑菌活性。     

Disposition of leucovorin and its metabolites in dietary folic acid-deplete mice – comparison between tumor, liver and plasma

Purpose: A comprehensive pharmacokinetic study of leucovorin (5-formyltetrahydrofolate, 5-HCO-FH₄) and its metabolites was conducted in plasma, liver and implanted tumor tissue from mice maintained on a low folic acid diet. While it has been previously demonstrated that the antitumor activity of fluorouracil (FU) can be potentiated by 5-HCO-FH₄, the optimum time for administration of FU after 5-HCO-FH₄, to maximally elevate the active folate metabolite methylenetetrahydrofolate in tumor has not been established. Human plasma studies have defined the pharmacokinetics of circulating 5-HCO-FH₄ and its metabolites, but comparison with human tumor accumulation has not been practicable because of sampling difficulties. As an alternative, a mouse model system, based on low dietary folic acid, was used to evaluate plasma, liver and implanted tumor reduced folates after administration of 5-HCO-FH₄.Methods: Plasma and tissue samples were collected from folate-deplete mice over a 12-h period after intraperitoneal administration of 90 mg/kg [R, S ] 5-HCO-FH₄. Reduced folates were evaluated using a ternary complex assay. Results: The time at which max‐imal accumulation of parent compound and all metabolites, except 5-methyltetrahydrofolate (5-CH₃FH₄), occurred in tumor was the same as in plasma. Alternatively, peak liver accumulation was delayed relative to plasma for all folates except 5-CH₃FH₄. Conclusions: The results suggest that mouse plasma accumulation of reduced folates, with the exception of 5-CH₃FH₄, can predict tumor accumulation. Hence, evaluation of human plasma folate accumulation may potentially provide a means to improve the timing of the administration of FU relative to 5-HCO-FH₄ to achieve a superior therapeutic outcome. Received: 24 July 1996 / Accepted: 29 October 1996     

宫颈癌组织中HPV16 E7序列多态性分析

目的探讨广东地区宫颈癌组织中HPV16肿瘤相关性抗原E7基因序列的多态性.方法采用通用引物PCR直接测序法对宫颈癌标本中的HPV分型,从含有HPV16型的标本中采用自行设计的多重引物通过巢式PCR扩增出HPV16E7,经DNA序列测定法检测其基因变异,进而寻找其热点突变.结果50例宫颈癌组织HPV-DNA的检出率为78%,其中HPV16和HPV18型混合感染18例,单纯HPV16型感染15例.33例含有HPV16型的标本中扩增出25例HPV16E7,其中17例647位核苷酸“T”变异“C”,导致相应的蛋白质由天冬氨酸变异为丝氨酸.结论广东地区宫颈癌组织中HPV16E7DNA序列发生碱基替换的区域主要在647位至846位,热点突变点为Nt647和Nt846.     

Measurement of contact impedance of electrodes used for deep brain stimulation

High frequency electrical stimulation by means of electrodes implanted into the brain has become an accepted technique for treatment of Parkinson's disease. The electrical field distribution normally inserted into the sub thalamic nucleus minimise abnormal brain activity. Square wave pulses of 1–3.6 V with duration of 60–90 μs at a frequency range of 130–185 pps are generally used. Every electrode unit consists of four cylindrical electrodes positioned in a row and can be switched on independently. This paper determines the contact impedance of the electrodes for different frequencies and proposes improvement to reduce the contact impedance between the electrodes and the brain. Measurements were performed by placing the electrodes in a tank filled with saline. Different frequencies were applied on two electrodes via a resistor. The current was measured through the resistor and the voltage was registered between one of the electrodes and a third non current carrying electrode. The obtained values were used to calculate the contact impedance. The result shows large contact impedance for the used frequency compared to the impedance of the treated tissue, which means that variation in contact impedance can result in variation in the electrical field applied to the tissue.