Altered copper level and renal dysfunction in Nigerian women using skin-whitening agents.

The purpose of the study was to determine the concentration of trace elements in serum samples of women who are chronic users of skin-whitening agents as well as the hepatic and renal effects of these agents on these women. The study was conducted among 23 skin-whitening users while 25 women served as the controls. The serum concentrations of Zn, Mn, and Mg were not significantly changed in these women compared with controls (p > 0.05), but Cu was significantly increased in skin-whitening agent users compared with controls (p < 0.05). Serum urea and creatinine were significantly increased compared with the controls (p < 0.05). Moreover, ALT, AST, albumin, total protein, and bilirubin were not significantly changed (p > 0.05). The significant increase in the levels of renal indices shows that these agents might be nephrotoxic after prolonged usage while increase in copper level with accompanying renal dysfunction may be an indication that copper mediates in oxidative-induced renal dysfunction. However, further study is needed to identify the cause and source of high serum copper as many of the herbal extracts may be rich sources of copper. Moreover, a large population study may be necessary to examine the exact correlation between copper and renal indices.

Cyclobutane-pyrimidine dimer excision in UV-sensitive CHO mutants and the effect of the human ERCC2 repair gene.

Using a radiochromatographic assay, we have examined cis-syn cyclobutane-pyrimidine dimer removal after ultraviolet irradiation in cell lines representative of the first 6 complementation groups of Chinese hamster ovary DNA nucleotide excision repair mutants. AA8, the CHO cell line from which these mutants were derived, consistently showed normal dimer excision for a rodent cell. The mutants uniformly exhibited no significant dimer excision within the limits of determination. Additionally, V-H1, a mutant belonging to complementation group 2 and derived from V79 hamster cells, exhibited no dimer excision. Two UV5 derived transformants that carry the complementing human ERCC2 repair gene showed a capacity for dimer excision comparable to the AA8 wild-type cells.

Detection and repair of a UV-induced photosensitive lesion in the DNA of human cells.

Irradiation with UV light results in damage to the DNA of human cells. The most numerous lesions are pyrimidine dimers; however, other lesions are known to occur and may contribute to the overall deleterious effect of UV irradiation. We have observed evidence of a UV-induced lesion other than pyrimidine dimers in the DNA of human cells by measuring DNA strand breaks induced by irradiating with 313-nm light following UV (254-nm) irradiation. These breaks, measured by alkaline sucrose sedimentation, increased linearly with the dose of UV light over the range tested (10-40 J/m2). The breaks cannot be photolytically induced 5 h after a UV dose of 20 J/m2 in normal cells; however, in xeroderma pigmentosum variant cells, the breaks are inducible for up to 24 h after UV irradiation. Xeroderma pigmentosum group A cells in the same 5-h period show an increase in the number of strand breaks seen with 313-nm light photolysis from about 2 to 4 breaks/10(9) dalton DNA. These breaks can then be induced for up to 24 h. These data suggest that, in normal cells, the lesion responsible for this effect is rapidly repaired or altered; whereas, in xeroderma pigmentosum variant cells it seems to remain unchanged. Some change apparently occurs in the DNA of xeroderma pigmentosum group A cells which results in an increase in photolability. These data indicate a deficiency in DNA repair of xeroderma pigmentosum variant cells as well as in xeroderma pigmentosum group A cells.

Effects of extremely low frequency (ELF) electric fields on cell growth and DNA repair in human skin fibroblasts.

A number of physical and chemical agents in the environment have been studied for their ability to induce or alter DNA repair mechanisms in human cells. We have investigated the effects of 60 Hz, 1000 V/cm electric fields on DNA repair in normal human fibroblasts in vitro. An examination was done on the ability of electric fields suspected to cause damage which could be repaired by thymine dimer excision and measurable by the bromodeoxyuridine photolysis assay. The thymine dimer assay with enzyme-sensitive site analysis was used to measure the cells' capacity for removing ultraviolet light (u.v.)-induced pyrimidine dimers; during exposure to electric field 24 hr before u.v. irradiation; 24 hr after u.v. irradiation; and up to 48 hr continuously after u.v. irradiation. Cell growth and cell survival following electric field exposure were also studied. Within the limits of these experiments, it was found that exposure to such electric fields did not alter cell growth or survival, and no DNA repair or alteration in DNA excision repair capacity was observed as compared with unexposed control cultures.

Selective potentiating effect of beta-p-chlorophenylglutamate on responses induced by certain sulphur-containing excitatory amino acids and quisqualate.

In dorsal horn neurones of the cat spinal cord iontophoretically administered (+/-)-beta-p-chlorophenylglutamate (chlorpheg) markedly enhanced the excitatory responses induced by L-homocysteate, L-homocysteine sulphinate, S-sulpho L-cysteine, L-cysteate and quisqualate, while responses to NMDA, kainate, L-glutamate, L-aspartate and L-cysteine sulphinate were generally unaffected. Preliminary data obtained on frog spinal cord in vitro supports the possibility that such selective potentiation may be due to differential inhibition by chlorpheg of amino acid uptake. No potentiating effects of chlorpheg were observed on spinal synaptic excitation.

Conformational aspects of the actions of some piperidine dicarboxylic acids at excitatory amino acid receptors in the mammalian and amphibian spinal cord.

A series of piperidine dicarboxylates (PDA) has been tested for excitatory amino acid agonist and antagonist activity and for synaptic depressant properties of the spinal cords of frogs and immature rats in vitro and of cats in vivo. The substances tested comprised (+/-)-cis-2,3-PDA, (+/-)-cis-2,4-PDA, (+/-)-cis-2,5-PDA, (+/-)-cis-2,6-PDA, (+/-)-trans-2,3-PDA, (+/-)-trans-2,4-PDA and both (+) and (-) forms of cis-2,3-PDA. Peak excitatory amino acid agonist activity was observed with (+/-)-trans-2,3- and (+/-)-trans-2,4-PDA. Excitatory amino acid antagonism and synaptic depressant activity was observed only with cis-dicarboxylates, this activity being greatest in the 2,3-analogue. The agonist actions of piperidine dicarboxylates were effectively depressed by the specific NMDA receptor antagonist, (-)-2-amino-5-phosphonovalerate and, where tested, also by D-alpha-aminoadipate and low concentrations of Mg2+. It was concluded that the major part of these agonist actions were mediated by NMDA receptors. The main structural feature of the NMDA agonist actions of these substances was considered to be their close relationship to N-alkyl-aspartic and glutamic acid molecules, with the trans arrangement of the respective 2,3- and 2,4-situated carboxyl groups promoting most effective interaction with the active sites of the NMDA receptor. (+/-)-Cis-2,3-PDA depressed excitatory responses induced by NMDA, kainate, quisqualate, (+/-)-trans-2,3-PDA and (+/-)-trans-2,4-PDA, or evoked by dorsal root stimulation. Both monosynaptic and polysynaptic excitation were susceptible to the depressant action of this substance. The (-) isomer of cis-2,3-PDA carried both excitatory amino acid agonist and antagonist activity and also the synaptic depressant properties observed with the racemic form of this substance. The (+) isomer showed little pharmacological activity. It is proposed that the structure-activity features of these heterocyclic amino acids indicate some of the conformational requirements for interaction with physiological excitatory amino acid receptors.

The effects of a series of omega-phosphonic alpha-carboxylic amino acids on electrically evoked and excitant amino acid-induced responses in isolated spinal cord preparations.

1 The depressant actions on evoked electrical activity and the excitant amino acid antagonist properties of a range of omega-phosphonic alpha-carboxylic amino acids have been investigated in the isolated spinal cord preparations of the frog or immature rat. 2 When tested on dorsal root-evoked ventral root potentials, members of the homologous series from 2- amino-5-phosphonovaleric acid to 2-amino-8-phosphonooctanoic acid showed depressant actions which correlated with the ability of the substances to antagonize selectivity motoneuronal depolarizations induced by N-methyl-D-aspartate. 3 2-Amino-5-phosphonovalerate was the most potent substance of the series giving an apparent KD of 1.4 microM for the antagonism of responses to N-methyl-D-aspartate. 4 A comparison of the (+)- and (-)-forms of 2-amino-5-phosphonovalerate indicated that the N-methyl-D-aspartate antagonist activity and the neuronal depressant action of this substance were both due mainly to the (-)-isomer. 5 The (-)- and (+)-forms of 2-amino-4-phosphonobutyrate had different actions. The (-)-forms of this substance had a relatively weak and non-selective antagonist action on depolarizations induced by N-methyl-D-aspartate, quisqualate and kainate and a similarly weak depressant effect when tested on evoked electrical activity. The (+)-form was more potent than he (-)-form in depressing electrically evoked activity but did not antagonize responses to amino acid excitants. At concentrations higher than those required to depress electrically evoked activity, the (+)-form produced depolarization. This action was blocked by 2-amino-5-phosphonovalerate.

Classification of chemical agents as to their ability to induce long- or short-patch DNA repair in human cells.

33 chemical agents and UV- and gamma-irradiation were tested for their comparative ability to induce long-patch or short-patch repair using the 5-bromodeoxy-uridine photolysis assay. For 11 chemical agents repair was long-patch in nature as determined by calculated patch size and response of xeroderma pigmentosum cells relative to normal human cells. Typical patch sizes as measured by this assay were about 90 nucleotides for UV repair, a range of 30 to 70 nucleotides for a variety of known and suspected UV-mimetic chemicals, and 3-4 nucleotides for gamma-radiation. Alkylating agents previously shown to induce short-patch repair were shown also to induce long-patch repair.

The relationship of DNA excision repair of ultraviolet-induced lesions to the maximum life span of mammals.

Physical and chemical agents present in the environment can potentially damage mammalian DNA. Such damage is known in some cases to be repaired by the process of DNA excision repair. This process has been extensively studied utilizing the repair of ultraviolet irradiation damage as a model system. In this study we have used this system and the 5-bromodeoxyuridine photolysis assay to measure DNA excision repair in cells derived from 21 mammalian species. We have attempted to relate the DNA repair proficiencies and the average size of the repaired regions seen in the cell cultures with the various maximum life spans of the mammals studied. There was an approximate linear correlation between life span of the mammals and the number of DNA excision repair sites measured 20-22 hours following ultraviolet irradiation of the cell cultures. Several deviations from the linear relationships were observed which remain largely unexplained. The size of the repaired regions was shown not to be related to the maximum life spans of the mammals tested.

Antagonism of excitatory amino acid-induced and synaptic excitation of spinal neurones by cis-2,3-piperidine dicarboxylate.

In tests on neurones in the cat spinal cord in vivo, and frog and immature rat spinal cord in vitro, cis-2,3-piperidine dicarboxylate (cis-2,3-PDA) produced the following effects: (1) selective antagonism of amino acid-induced responses, compared with responses to other putative transmitters; (2) effective antagonism of kainate and quisqualate-induced responses in addition to responses induced by N-methyl-D-aspartate (NMDA) and other excitatory amino acids; (3) partial NMDA-like agonist action; (4) antagonism of dorsal root-evoked excitation of Renshaw cells; (5) potentiation of acetylcholine- and ventral root-evoked excitation of Renshaw cells. This unique spectrum of action may be useful for transmitter receptor characterization in the vertebrate central nervous system.

2-Amino-5-phosphonovalerate (2APV), a potent and selective antagonist of amino acid-induced and synaptic excitation.

A new compound, 2-amino-5-phosphonovaleric acid (2APV) is the most potent and selective N-methyl-D-aspartate (NMDA) receptor antagonist yet tested. As with other compounds of this type, it blocks L-aspartate and dorsal root-evoked excitation of spinal neurons, but is without effect on the cholinergic excitation of Renshaw cells evoked by exogenous acetylcholine or ventral root stimulation. The high potency and selectivity of this compound should prove to be of great value in investigations of the amino acid receptor types involved in synaptic excitation.

Dipeptide antagonists of amino acid-induced and synaptic excitation in the frog spinal cord.

The dipeptide gamma-D-glutamylglycine (gamma DGG) antagonizes amino acid-induced depolarization and synaptic excitation in the isolated hemisected spinal cord of the frog. In general, the effects of this compound resembled those of the structurally similar D-alpha-aminosuberate (D alpha AS) in being more effective against N-methyl-D-aspartate (NMDA)-induced responses than against responses induced by other excitatory amino acids. However gamma DGG appeared to be more effective than D alpha AS in depressing kainate-induced responses. Similar, though weaker, effects were produced by the L isomer of the dipeptide (gamma LGG), a natural brain constituent.

Selective depression of excitatory amino acid induced depolarizations by magnesium ions in isolated spinal cord preparations.

1. The depressant actions of Mg2+ and a range of other divalent ions on synaptic excitation and on responses produced by excitatory amino acids and other putative transmitters have been investigated in hemisected isolated spinal cords of frogs and neonatal rats. Some comparative studies were also made using the rat isolated superior cervical ganglion. 2. At concentrations above 10 microM, Mg2+ selectively antagonized N-methyl-D-aspartate (NMDA)-induced motoneurone depolarization as recorded from ventral roots of tetrodotoxin-blocked spinal cords. Depolarization evoked by quisqualate (unaffected by 20 mM-Mg2+) was resistant to the depressant action of these ions, while depolarizations evoked by other excitant amino acids were depressed to intermediate degrees. 3. Mn2+, Co2+ and Ni2+ had qualitatively similar actions to Mg2+; Mn2+ was somewhat less potent and Co2+ and Ni2+ more potent than Mg2+. The alkaline earth metal ions, Ca2+, Sr2+ and Ba2+, had very weak Mg2+-like actions. Ca2+ and Mg2+ acted additively in depressing amino acid-induced responses. 4. Mg2+ also depressed motoneurone responses evoked by noradrenaline, substance P and carbachol in the neonatal rat isolated spinal cord. However, none of these effects were as marked as the depression of NMDA-induced responses by Mg2+ in this preparation. Mg2+ did not depress motoneurone depolarization produced by 5-HT in the rat spinal cord or the depolarizing action of GABA on primary afferent terminals of the isolated frog spinal cord. 5. At concentrations producing marked depression of NMDA-induced responses, Mg2+ also depressed synaptic transmission in spinal cords in the absence of an effect on ganglionic transmission. At the same concentrations, Mn2+, Co2+ and Ni2+ depressed synaptic transmission in both preparations. 6. From the similarity in action between Mg2+ and the D-alpha-aminoadipate group of NMDA antagonists, it is suggested that the central depressant action of low concentrations of Mg2+ involves predominantly a postsynaptically mediated interference with the action of an excitatory amino acid transmitter.

Antagonism of excitatory amino acid-induced responses and of synaptic excitation in the isolated spinal cord of the frog.

1. A range of compounds has been tested for excitatory amino acid agonist or antagonist activity and for effects on synaptic activity on isolated hemisected spinal cords of frogs. 2. L-Monoamino dicarboxylic acids of chain length up to 8 carbon atoms (L-alpha-aminosuberate) were all agonists. 3. Within a series of D-monoamino dicarboxylic acids, and with diamino dicarboxylic acids (mainly unresolved mixtures of diasteroisomers), there was a progression from agonist activity, for compounds of chain length equal to or shorter than glutamate, to antagonist activity, for compounds of longer chain length equal to or shorter than glutamate, to antagonist activity, for compounds of longer chain length, D-alpha-Aminosuberate (D alpha SD) was the most potent antagonist. 4. The antagonist actions of these substances showed a Mg2+--like selectivity with respect to depolarizations produced by different excitants. N-methyl-D-aspartate (NMDA) was the most susceptible agonist and quisqualate and kainate the least susceptible. Responses to other excitatory amino acids, including L-glutamate and L-aspartate, showed intermediate sensitivity to the antagonists. 5. A parallelism was observed between the relative potencies of mono- and diamino dicarboxylic acids as NMDA antagonists and their relative potencies as depressants of synaptic responses. 6. The results support the concept of different types of excitatory amino acid receptors, with NMDA and its antagonists acting predominantly on one type. These NMDA receptors are probably transmitter receptors activated by an excitatory amino acid transmitter.

Inhibition of DNA repair in ultraviolet-irradiated human cells by hydroxyurea.

The effect on DNA repair in ultraviolet-irradiated human skin fibroblasts by hydroxyurea has been examined in this study using three independent methods for measuring DNA repair:the 5-bromodeoxyuridine photolysis assay which measures DNA repair replication, chromatographic measurement of thymine-containing dimers, and measurement of specific ultraviolet-endonuclease-sensitive sites in irradiated DNA. Little effect of hydroxyurea was observed at the concentration of 2 mM, which is often used to inhibit semiconservative DNA synthesis; however, 10 mM hydroxyurea resulted in marked inhibition (65--70%) of excision repair. This inhibition was accompanied by a possible doubling in the size of the repaired region. The accumulation of large numbers of single-strand breaks following ultraviolet irradiation and hydroxyurea incubation seen by other investigators was not observed with the normal skin fibroblasts used in this study. A comparison of hydroxyurea effects on the different DNA repair assays indicates inhibition of one step in DNA repair also results in varying degrees of inhibition of other steps as well.

Excitatory amino acid receptors and synaptic excitation in the mammalian central nervous system.

At least two different types of excitatory amino acid receptors have been identified in the mammalian and amphibian central nervous systems. One type ('NMDA receptors') appears to be important in amino acid-mediated synaptic excitation, NMDA being the most potent and specific exogenous agonist for this type of receptor. Many antagonists have selective blocking actions at these NMDA receptors, and such substances are also selective antagonists of synaptic excitation in the vertebrate spinal cord. It is proposed that these receptors are transmitter receptors activated by an excitatory amino acid. In addition, extrasynaptic receptors, activated by domoate, kainate, quisqualate and L-glutamate, but not by NMDA, and only weakly by L-aspartate, have been identified on dorsal root fibres of the immature rat.

Quisqualamine, a novel gamma-aminobutyric acid (GABA) related depressant amino acid.

A new substnace, quisqualamine, the decarboxylated analogue of quisqualic acid, predictably depressed electrical activity of neurons of the frog and rat spinal cord in vitro and of the mouse spinal cord in vivo. In the in vitro preparations, the action of quisqualamine was associated with a prolonged depolarization of primary afferent terminals which was sensitive to blockade by picrotoxin and bicuculline and which was also depressed by strychnine. This suggests an interaction of quisqualamine with presynaptic receptors for both GABA and beta-alanine. Post-synaptic actions of quisqualamine, which were less marked than those at presynaptic sites, also appeared to be predominantly GABA-mimetic in vitro, though a sensitivity to the GABA-antagonist bicuculline could not be demonstrated in vitro.