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Summary Background Blue-collar workers have a high risk of occupational contact dermatitis, but epidemiological studies are scarce. Objectives To investigate allergic contact dermatitis in blue-collar workers with dermatitis registered by the Danish Contact Dermatitis Group. Methods A retrospective analysis of patch test data from 1471 blue-collar workers and 1471 matched controls tested between 2003 and 2012 was performed. A logistic regression was used to test for associations. Results The blue-collar workers often had occupational hand dermatitis (p < 0.001). Atopic dermatitis was less commonly observed among blue-collar workers (19.6%) than among controls (23.9%) (p = 0.005). Allergens with a statistically significant association with the occupational group of blue-collar workers were epoxy resins, methyldibromo glutaronitrile, 2-bromo-2-nitro-1,3-propanediol, potassium dichromate, and methylchloroisothiazolinone (MCI)/methylisothiazolinone (MI). The following occupations were additionally identified as risk factors for contact sensitization to MCI/MI and MI, epoxy resins, and potassium dichromate, respectively: painting, construction work, and tile setting/terrazzo work. Conclusion Contact allergy is a major problem among blue-collar workers. The data indicate a healthy worker effect among blue-collar workers diagnosed with dermatitis, as blue-collar workers were diagnosed significantly less often with atopic dermatitis than were controls.

SummaryBackground Occupational contact dermatitis among hairdressers is frequent, owing to daily exposure to irritants and allergens. Objectives To identify sensitization to the most common allergens associated with the occupation of hairdressing. Methods Patch test results of 399 hairdressers and 1995 matched controls with contact dermatitis, registered by the Danish Contact Dermatitis Group between January 2002 and December 2011, were analysed. All patients were patch tested with the European baseline series, and hairdressers were additionally tested with the hairdressing series. Results Occupational contact dermatitis (p

Background. No generally accepted classification scheme for hand eczema exists. The Danish Contact Dermatitis Group recently developed a guideline defining common clinical types and providing criteria for aetiological types. Objectives. To test the concepts of this guideline in a group of hand eczema patients. Methods. Seven hundred and ten hand eczema patients were included from seven dermatology clinics in Denmark. The hand eczema was classified into one of five clinical types, with standard photographs as reference. The severity was scored by the physician, who also made a final aetiological diagnosis. Results. Irritant contact dermatitis was most frequent in chronic, dry fissured hand eczema (44.3%), pulpitis (41.7%), and nummular hand eczema (40.9%), whereas allergic contact dermatitis dominated in vesicular types of hand eczema, with recurrent (35%) and few (24.2%) eruptions. Hyperkeratotic palmar hand eczema was the only clinical type that constituted a distinct subgroup; it was found most frequently in older men and had the strongest relationship, although not significant, with non-specific dermatitis. Conclusions. The relationship between clinical type of hand eczema and aetiological diagnosis fitted with general experience, but no simple relationship was found. This emphasizes that patch testing and exposure analysis are mandatory. Hyperkeratotic palmar hand eczema was identified as a distinct clinical subtype.

Udgivelsesdato: 2008-Jun-4 Background: The preservative methyldibromo glutaronitrile (MDBGN) has been banned, first from stay-on, and later from rinse-off cosmetics, in the EU countries because of increasing rates of contact allergy. Objectives: To evaluate the frequency of contact allergy to MDBGN among patients patch tested by the Danish Contact Dermatitis Group just before and following regulatory decisions. Patients/Methods: The data set comprised 19 279 consecutive eczema patients patch tested from 2003-2007 with MDBGN 0.3% in petrolatum (pet.) or, in a minority of patients, with Euxyl Ktrade mark 400 1.5% in pet. Results: A significantly decreasing trend in the frequency of positive patch tests to MDBGN was found from 4.6% in 2003 to 2.6% in 2007 (P < 0.001). The decreasing trend was seen for both men and women. A significantly decreasing proportion of cases with a current relevance of contact allergy to MDBGN was also seen from 51.3% in 2003 to 29% in 2007 (P < 0.001). Conclusions: Regulatory interventions have already had a major effect on allergic disease due to MDBGN in Denmark. The same trends are likely to be seen in other EU countries.

The homogeneity of the distribution of the non-selective voltage-dependent cation channel (the NSVDC channel) in the human erythrocyte, and the pH dependence was investigated. Activation of this channel caused a uniform cellular dehydration, which was characterized by the changes in the erythrocyte osmotic resistance profiles: after 1/2 h of activation, the osmolarity at 50% hemolysis changed from 73 mM (control) to 34 mM NaCl, corresponding to 0.48% and 0.21% NaCl respectively. Unchanging standard deviations show participation of the entire erythrocyte population, which implies an even distribution of the NSVDC channel among the cells. Inactivation of the NSVDC channel with N-ethyl-maleimide (NEM) or blocking of the Cl(-) conductance with NS1652 retarded the migration of the resistance profiles towards lower osmolarities. The NSVDC channel activation was blocked by a decrease of the intracellular -- but not the extracellular -- pH. The apparent pK(A) value for the effect was estimated to be 6.5, and the specific histidine reagent 2.4'-dibromoacetophenone (DBAB) inactivated the NSVDC channel.

Pharmacological modulation of the nonselective voltage-dependent cation (NSVDC) channel from human erythrocytes was studied. Using the inorganic cations ruthenium red and La3+, as well as the organic thiol group reagents iodoacetamide (IAA) and N-ethylmaleimide (NEM), it was possible to demonstrate a concentration-dependent decrease in the voltage-activated conductance, reflecting an inhibition or inactivation of the channel. Initial voltage activation was achieved by injecting human red cells into sucrose-substituted Ringers with a low chloride concentration, which causes a strongly positive membrane potential to develop, initially determined by the equilibrium potential for Cl- ( approximately +100 mV). Due to the voltage- and time-dependent activation of the cation channel, net effluxes, minimized by addition of a chloride conductance blocker, occurred and Vm gradually decreased and stabilized at a value less positive than E(Cl), reflecting the increased cation conductance, g+, reaching 1.5-2.0 microS/cm2. In the presence of inhibitors of the NSVDC channel, both the membrane potential repolarization and the cation efflux were diminished.

The activation and pharmacological modulation of the nonselective voltage-dependent cation (NSVDC) channel from human erythrocytes were studied. Basic channel activation was achieved by suspending red cells in a low Cl − Ringer (2 mM), where a positive membrane potential ( V m = E Cl ) immediately developed. Voltage- and time-dependent activation of the NSVDC channel occurred, reaching a cation conductance ( g + ) of 1.5–2.0 μS cm −2 . In the presence of the classical Gardos channel blocker clotrimazole (0–50 μM), activation occurred faster, and g + saturated dose-dependently (EC 50 = 14 μM) at a value of about 4 μS cm −2 . The clotrimazole analogues TRAM-34, econazole, and miconazole also stimulated the channel, whereas the chemically more distant Gardos channel inhibitors nitrendipine and cetiedil had no effects. Although the potency for modulation of the NSVDC channel is much lower than the IC 50 value for Gardos channel inhibition, clotrimazole (and its analogues) constitutes the first chemical class of positive modulators of the NSVDC channel. This may be an important pharmacological “fingerprint” in the identification of the cloned equivalent of the erythrocyte channel.

The activation/deactivation kinetics of the human erythrocyte voltage-dependent cation channel was characterized at the single-channel level using inside-out patches. It was found that the time dependence for voltage activation after steps to positive membrane potentials was slow ( t(1/2) about 30 s), whereas the deactivation was fast ( t(1/2) about 15 ms). Both activation and deactivation of this channel were also demonstrated in intact red cells in suspension. At very positive membrane potentials generated by suspension in extracellular low Cl(-) concentrations, the cation conductance switched on with a time constant of about 2 min. Deactivation of the cation channel was clearly demonstrated during transient activation of the Gárdos channel elicited by Ca(2+) influx via the cation channel and ensuing efflux via the Ca(2+) pump. Thus, the voltage-dependent cation channel, the Gárdos channel and the Ca(2+) pump constitute a coupled feedback-regulated system that may become operative under physiological conditions.

detergents; hydroxyisohexyl-3-cyclohexene carboxaldehyde; irritant contact dermatitis; cosmetics.Hydroxyisohexyl 3-cyclohexene carboxaldehyde (HICC)is a fragrance ingredient that is often used in cosmeticproducts (1, 2). Because of this frequent use in cosmeticproducts, as well as its use in higher concentrationsthan the recommended safe-use concentration, HICCis one of the most frequent causes of allergic contactdermatitis (3–10). In order to reduce the prevalenceof contact allergy to HICC, the International Fragrance

In order to examine whether calcium-dependent binding of annexin to acidic phospholipids could change the lipid bilayer environment sufficiently to perturb channel-mediated transmembrane ion-transport, gramicidin A channel activity in planar lipid bilayers was investigated in the presence of calcium and annexins II, III or V. The experiments were performed with membranes consisting of phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine in 300 mM KCl solution buffered to pH 7.4 and with either 0.1 or 1 mM calcium added to the solution. Annexin (1 microM) was subsequently applied to the cis side of the membrane. All three annexins (II, III and V) when tested at 1 mM calcium decreased the gramicidin single-channel conductance. Annexins II and III increased the mean lifetime of the channels whereas annexin V seemed to have no influence on the mean lifetime. Since the lifetime of gramicidin A channels is a function of the rate constant for dissociation of the gramicidin dimer, which is dependent on the physical properties of the lipid phase, binding of annexins II and III seems to stabilize the gramicidin channel owing to a change of the bilayer structure.

We have isolated calpromotin, a protein reported abundant in human red cells and shown to be of significance for KCl transport. We show that calpromotin is identical to a radical scavenger protein, thiol-specific antioxidant protein (TSA). Calpromotin is known to exist partially as a large complex of identical subunits and partially as dimers probably held together by disulfide bridges. The stability of the high-molecular-weight form was studied by variations of pH and urea concentration. It is shown that the equilibrium between the large complex and the dimeric subunit is governed by the dissociation of a group with a pK value of about 7.5. Dissociation of the complex was also complete at 2.5 M urea, where no unfolding of the peptide chains was detectable.

The effect of annexins II, III and V, purified from different species, on the calcium-activated chloride current across the stage-V to stage-VI Xenopus laevis oocyte membrane was tested either directly, using calcium entry mediated by depolarization, by A23187 permeabilization of oocytes or indirectly by quisqualate stimulation of a metabotropic glutamate receptor in the membrane expressed by the oocyte after injection of mRNA. The annexins isolated from the Ehrlich ascites cell, which is a mouse tumor cell, were found to be potent inhibitors of the chloride current, showing half-maximal inhibition at 50 nM, whereas no block was found using bovine or porcine annexins isolated from lung tissue. Of the annexins tested, we found annexin III to be naturally occurring in the oocyte, while only trace amounts of annexins II and V could be demonstrated. The inhibition pattern varied somewhat according to the stimulus method, the inhibition being more complete when an indirect stimulus via the metabotropic receptor was applied compared to a direct calcium stimulus.

Background. Painters are among the occupational groups that most commonly experience occupational contact dermatitis, but few investigations exist concerning this occupation. Objectives. To characterize painters with contact dermatitis and identify the most common allergens associated with the occupation. Materials and methods. All patch test results of 219 painters and 1095 matched controls registered by the Danish Contact Dermatitis Group between 2001 and 2010 were analysed. Results. Hand eczema (p < 0.0001) and occupational contact dermatitis (p < 0.0001) were observed significantly more often in the painters than in the group of controls. Sensitizations to the following allergens from the European baseline series were associated with the occupation and were statistically significant: methylchloroisothiazolinone/methylisothiazolinone, epoxy resin, formaldehyde, and quaternium-15. Three different isothiazolinones emerged as the most frequent sensitizers of the allergens tested in addition to the baseline series. Conclusions. The results indicate that painters have an increased risk of developing occupational hand eczema. Isothiazolinones and epoxy resin proved to be the two most frequent sensitizers in painters.

Background. Epoxy resin monomers are strong skin sensitizers that are widely used in industrial sectors. In Denmark, the law stipulates that workers must undergo a course on safe handling of epoxy resins prior to occupational exposure, but the effectiveness of this initiative is largely unknown. Objectives. To evaluate the prevalence of contact allergy to epoxy resin monomer (diglycidyl ether of bisphenol A; MW 340) among patients with suspected contact dermatitis and relate this to occupation and work-related consequences. Patients/methods. The dataset comprised 20 808 consecutive dermatitis patients patch tested during 2005-2009. All patients with an epoxy resin-positive patch test were sent a questionnaire. Results. A positive patch test reaction to epoxy resin was found in 275 patients (1.3%), with a higher proportion in men (1.9%) than in women (1.0%). The prevalence of sensitization to epoxy resin remained stable over the study period. Of the patients with an epoxy resin-positive patch test, 71% returned a questionnaire; 95 patients had worked with epoxy resin in the occupational setting, and, of these, one-third did not use protective gloves and only 50.5% (48) had participated in an educational programme. Conclusion. The 1% prevalence of epoxy resin contact allergy is equivalent to reports from other countries. The high occurrence of epoxy resin exposure at work, and the limited use of protective measures, indicate that reinforcement of the law is required.

Background. Fragrances frequently cause contact allergy, and cosmetic products are the main causes of fragrance contact allergy. As the various products have distinctive forms of application and composition of ingredients, some product groups are potentially more likely to play a part in allergic reactions than others. Aim. To determine which cosmetic product groups cause fragrance allergy among Danish eczema patients. Method. This was a retrospective study based on data collected by members of the Danish Contact Dermatitis Group. Participants (N = 17716) were consecutively patch tested with fragrance markers from the European baseline series (2005–2009). Results. Of the participants, 10.1% had fragrance allergy, of which 42.1% was caused by a cosmetic product: deodorants accounted for 25%, and scented lotions 24.4%. A sex difference was apparent, as deodorants were significantly more likely to be listed as the cause of fragrance allergy in men (odds ratio 2.2) than in women. Correlation was observed between deodorants listed as the cause of allergy and allergy detected with fragrance mix II (FM II) and hydroxyisohexyl 3-cyclohexene carboxaldehyde. Conclusion. Deodorants were the leading causes of fragrance allergy, especially among men. Seemingly, deodorants have an ‘unhealthy’ composition of the fragrance chemicals present in FM II.

The basal uptake and cytoplasmic binding of cobalt was studied in human red cells using (57)Co as tracer. The basal uptake is linear with time, at a rate of about 10 μmol (l cells)(-1) h(-1) at 100 μM [Co(2+)](o), and is almost irreversible, as there is hardly any efflux into excess EDTA. Ionophore A23187 mediates a rapid equilibration of Co(2+) across the cell membrane leading to a marked accumulation, reflecting effective cytoplasmic buffering. The fraction (α(Co)) of total cell cobalt being present as free, ionized Co(2+) is estimated at α(Co)=0.01 from the equilibrium distribution of cobalt, and also from the initial slope of the cobalt buffering curve. The cobalt accumulation is similar in fed and ATP-depleted cells. The buffering curve for [Co(T)](c) can be fitted by a Michaelis type function with B(max)=24 mmol (l cells)(-1) and half-saturation at 240 μM [Co(2+)](c). The tracer influx curves are adequately fitted by single exponentials, whereas the net influx curves all require at least double exponential fits, probably due to non-stationary A23187 kinetics. The rate of tracer influx decreases with increasing cobalt concentration, and increases with delayed addition of (57)Co tracer during net uptake. This might be explained by an 'auto-inhibition' by cobalt. The kinetics for A23187-mediated net and tracer influx of (54)Mn is very similar to that of (57)Co, whereas the net influx of (65)Zn can be fitted by single exponentials. In cobalt-loaded cells the cobalt is partly reversibly bound, being releasable by excess extracellular EGTA in the presence of A23187, and partly tightly bound, remaining in the cells even at high ionophore concentrations. The tightly bound fraction builds up over time, and is larger and develops earlier in fed cells compared to ATP-depleted cells. However, all cell cobalt appears to exchange with (57)Co during tracer influx. It is speculated that oxidation of Co(2+) to Co(3+) could lead to the high affinity binding. Tight binding is not observed in the case of (54)Mn. Tightly bound and the major part of reversibly bound (57)Co co-migrate with hemoglobin in Sephadex column chromatography of a lysate of (57)Co-loaded cells. (57)Co also co-migrates with hemoglobin when added to a lysate of unlabeled cells or to a solution of purified hemoglobin, in both cases with a time-dependent development of tight binding. Cobalt is known to bind to the globin moiety of hemoglobin. The results imply that during long-term cobalt exposure in vivo cobalt will be taken up practically irreversibly in the red cells during their 120 days life span. Thus, for biomonitoring of cobalt exposure, it could be appropriate to measure the cobalt content in red cells to give, compared with timed or in-competition whole-blood and serum analysis, an average value for the exposure over the last couple of months.

1. 1. Three calcium-binding proteins have been purified from Ehrlich ascites tumor cells. 2. 2. They were identified by amino acid sequence analysis on selected fragments obtained by tryptic digestion. 3. 3. The proteins belong to the annexin family and were identified as annexins II, III and V. 4. 4. Antibodies raised against the proteins were used to examine for their presence in a number of murine tissues. 5. 5. The occurrence was found to be in reasonable accordance with earlier reports.

Background: Fragrance mix II (FM II) is a relatively new screening marker for fragrance contact allergy. It was introduced in the patch test baseline series in Denmark in 2005 and contains six different fragrance chemicals commonly present in cosmetic products and which are known allergens. Aim: To investigate the diagnostic contribution of including FM II in the baseline series by comparing it with other screening markers of fragrance allergy: fragrance mix I (FM I), Myroxylon pereirae and hydroxyisohexyl 3-cyclohexene carboxaldehyde (HICC). Method: Retrospective study of 12 302 patients consecutively patch tested with FM II by members of the Danish Contact Dermatitis Group 2005–2008. Results: FM II gave a positive patch test in 553 patients (4.5%), and in 72.2% of these patients the reaction was judged to be clinically relevant. FM II ranked second in detecting fragrance allergy, after FM I. If FM II had not been included as a screening marker in the baseline series, 15.6% (n = 202) of individuals with fragrance allergy would not have been identified by the other fragrance screening markers (FM I, M. pereirae or HICC). Conclusion: FM II contributes substantially to detecting fragrance allergy. It ranked second among the fragrance screening markers tested in the baseline series and detects individuals with an allergy who otherwise would not have been identified.

Udgivelsesdato: 2009-Jun BACKGROUND: Hand eczema (HE) is a frequent, long-lasting disease with both personal and societal repercussions. Consequently, more information is needed on factors that maintain symptoms. OBJECTIVES: In this study, patients with HE were followed for 6 months from the first visit to a dermatologist to identify factors associated with severe disease and a poor prognosis. METHODS: Study participants were 799 patients with HE from nine dermatological clinics in Denmark. Severity assessment of the HE was done at baseline and at the 6-month follow-up using the Hand Eczema Severity Index (HECSI) and by patients using a self-administered photographic guide. Additional information was obtained from a baseline questionnaire. RESULTS: At baseline, 60.3% assessed their HE as moderate to very severe using the self-administered photographic guide compared with 36.1% at follow-up. The mean HECSI value decreased from 19.9 points at baseline to 11.2 points at follow-up (P < 0.001). In a multivariable logistic regression analysis, statistically significant associations with severe HE at baseline were older age (P < 0.001), atopic dermatitis (P = 0.01) and > or = 1 positive patch test (P < 0.001). Being an unskilled worker was a predictor for a poor prognosis at follow-up (P = 0.04), and the presence of frequent symptoms during the previous 12 months was associated with severe initial disease (P = 0.02) and a poor prognosis (P = 0.04). CONCLUSIONS: Overall, the disease had improved 6 months after the dermatological examination: nevertheless, many patients continued to have significant symptoms. Dermatologists should pay special attention to patients with frequent eruptions and to unskilled workers.

The activation and pharmacological modulation of the nonselective voltage-dependent cation (NSVDC) channel from human erythrocytes were studied. Basic channel activation was achieved by suspending red cells in a low Cl(-) Ringer (2 mM), where a positive membrane potential (V(m) = E(Cl)) immediately developed. Voltage- and time-dependent activation of the NSVDC channel occurred, reaching a cation conductance (g+) of 1.5-2.0 microS cm(-2). In the presence of the classical Gárdos channel blocker clotrimazole (0-50 microM), activation occurred faster, and g+ saturated dose-dependently (EC50 = 14 microM) at a value of about 4 microS cm(-2). The clotrimazole analogues TRAM-34, econazole, and miconazole also stimulated the channel, whereas the chemically more distant Gárdos channel inhibitors nitrendipine and cetiedil had no effects. Although the potency for modulation of the NSVDC channel is much lower than the IC50 value for Gárdos channel inhibition, clotrimazole (and its analogues) constitutes the first chemical class of positive modulators of the NSVDC channel. This may be an important pharmacological "fingerprint" in the identification of the cloned equivalent of the erythrocyte channel.

Suspension of intact human red cells in media with low chloride and sodium concentrations (isotonic sucrose substitution) results in strongly inside positive membrane potentials, which activate the voltage-dependent non-selective cation (NSVDC) channel. By systematic variation of the initial Nernst potentials for chloride (degree of ion substitution) as well as the chloride conductance (block by NS1652), and by exploiting the interplay between the Ca(2+)-permeable NSVDC channel, the Ca(2+)-activated K+ channel (the Gardos channel) and the Ca(2+)-pump, a graded activation of the NSVDC channel was achieved. Under these conditions, it was shown that the NSVDC channels exist in two states of activation depending on the initial conditions for the activation. The hysteretic behaviour, which in patch clamp experiments has been found for the individual channel unit, is thus retained at the cellular level and can be demonstrated with red cells in suspension.

1. 1. A long-term learning phenomenon in the tentacle of Helix pomatia has been observed. 2. 2. Repeated mechanical stimulation of the optic tentacle led to habituation of the associated withdrawal-extension action pattern whereas repeated combined mechanical and electrical stimulation potentiated the reflex. 3. 3. The potentiation was present 40 days after four sessions of stimulations. Sessions spaced with proportionally increasing intervals were more effective than massed training. Puromycin (125 mg/ml) blocked 80% of protein synthesis as well as the long-term but not the short-term retention of potentiation. 4. 4. Contrary to observations in other preparations, the amount of water soluble proteins, notably S-100, was not affected by learning. Injections in vivo of and exposure of brain in vitro to S-100 had no effect on learning or a variety of brain potentials.

The role of calmodulin in stimulating active calcium transport in the human red cell membrane is well documented. In contrast, efforts to characterize the effect of calmodulin on the Ca2+-dependent K+ channel in erythrocyte membranes have given rise to conflicting reports. These studies have indicated that experimental conditions may play a critical role in preserving the Ca2+-dependent K+ channels in erythrocyte inside-out vesicles. With these observations in mind, a double-labelling study of simultaneous active Ca2+ and passive Rb+ uptake in red-cell inside-out vesicles was undertaken. Addition of calmodulin and ATP to a suspension of inside-out vesicles containing 1 mM K+ caused a Ca2+-dependent increase in both the rate of active calcium transport and Rb+ uptake. The initial Rb+ isotope flux was increased 3-fold over the rate observed in the absence of calmodulin. The k 1 2 for activation of K+ permeability was approx. 5 · 10−7 M Ca2+ as compared to 10−6 M Ca2+ for active Ca2+ transport. Addition of the calmodulin antagonists pimozide and chlorpromazine blocked calmodulin activation of the Ca2+-dependent K+ channel. The observation that activation of the K+ channel occurs at Ca2+ concentrations which are lower than those required for maximum stimulation of the calcium pump suggests that these processes are dependent on two states of the calmodulin molecule, characterized by a lower or higher amount of Ca2+ bound to calmodulin.

Two to 3 days after the final ecdysis adult desert locusts were injected with a single dose of tritium-labelled tyrosine and its incorporation into resilin and resilin-secreting cells was studied as a function of time by means of autoradiography. The cells appear to be uniformly labelled after only 7 min. After 20 to 30 min there is a concentration of grains at the apical border and an indication of label in the newly secreted resilin. After 45 min the secretion of resilin reaches a peak. After 5 hr the cells and therefore the haemolymph are almost free of radioactivity.

By electron microscopy, the prominent bundles of filaments occurring in the basal part of proximal and distal tubule cells and in interstitial cells of rat kidney cortex were studied in cells fixed by vascular perfusion, in glycerol-extracted cells and in glycerol-extracted cells treated with heavy meromyosin (HMM). The studies of perfusion-fixed tissue showed that the proximal tubule cells contained in their most basal part filamentous bundles oriented transversely around the tubule. The bundles consisted of trightly packed thin filaments (50–80 A in diameter). Similar but less prominent bundles were found in distal tubule cells and in interstitial cells. The dimension of these filaments was similar to that of actin filaments and their insertion in the basal cell membrane of the tubule epithelial cells resembled the insertion of actin filaments in the cell membrane of smooth muscle cells. The studies on glycerol-extracted cells revealed that some tubule cells contained two types of filaments (60–80 A and 130–170 A in diameter) located side by side in the basal filamentous bundles. The dimension of the thick filaments corresponds well to the values for myosin filaments in glycerinated smooth and skeletal muscle. The studies on HMM-reacted renal tissue revealed that the thin filaments (60–80 A) described in tubule and interstitial cells are probably actin filaments, as they formed characteristic arrowhead complexes morphologically indistinguishable from the complexes of HMM with actin filaments in smooth and striated muscle cells. Our results provide strong evidence that a two-filament contractile system, based on interaction of actin and myosin filaments, exists in renal tubule and interstitial cells. As a hypothesis it is proposed that it is changes in tonus of the basal filamentous system in the proximal tubule cells which stabilize the intratubular pressure, possibly via angiotensin.

Incorporation of l -tyrosine, generally labelled with tritium and given as a single injection into the haemolymph, is shown to take less than 6 hr, and the lag-phase is less than 1–2 hr. The brightly fluorescent bands in resilin are synthesized during the day periods, whereas the faintly fluorescent bands are synthesized during the night periods. It is shown that the part of the prealar arm where two chitin lamellae are missing is synthesized soon after emergence.

Calmodulin binds to inside-out vesicles of the human red cell membrane in a Ca2+-dependent manner. Mg2+ was found to be essential to this binding, half maximum binding occurring at decreasing Ca2+ concentration with increasing Mg2+ concentration. The calmodulin antagonist calmidazolium strongly reduced the Ca2+-dependent binding of calmodulin to the inside-out vesicles.

Red cells from the giant salamander Amphiuma means are shown to contain sialic acid. The amount removed by the action of neuraminidase is equal to that released by acid hydrolysis, indicating that all of the sialic acid is present on the outer surface of the plasma membrane. These cells have a negative electrophoretic mobility and 100% enzymatic removal of sialic acid results in a 40% reduction in the mobility, suggesting that either a fraction of the sialic acid carboxyl groups are unavailable to the action of external electric fields, or other negatively charged groups contribute to the surface charge. A further reduction in mobility of normal and sialic acid-free cells is caused by an increased extracellular calcium concentration. The negative groups affected by calcium are most likely to be phosphate groups, since the isoelectric point of the cells is found to lie between the p K values for H2PO4− groups and the carboxyl groups of sialic acid. Membrane potentials of single cells, from which 80% or more of the total sialic acid had been removed, were identical to those measured in normal cells, confirming that sialic acid plays little, if any, direct role in the maintenance of membrane potentials and ionic permeabilities.

A set of proteins in the 33–37 kDa range have been isolated from the cytoplasm of the Ehrlich ascites tumor cell. The proteins are characterized by their Ca2+-dependent binding to cell membranes. This property has been used for isolation of the proteins by Ca2+-dependent affinity binding to inside-out vesicles of the human red cell membrane. The proteins display Ca2+-binding properties as shown by gel-filtration studies. The Ca2+-dependent binding of the 33 and 34 kDa proteins to red cell membranes was studied after labelling of the proteins with tritium by reductive methylation. The average number of Ca2+ bound per protein molecule was 4.8 with a Kd of 3.4 · 10−4 M Ca2+. The proteins are distinct from most other Ca2+-binding proteins of comparable molecular weights by not incorporating phosphate.

The structure of glycerinated smooth muscle from small intestine of adult rat was investigated by electron microscopy. In the central parts of the tissue blocks a two-filament system was found, consisting of parallel thick and thin filaments with regularly spaced interconnections, closely resembling that of striated muscle. In the peripheral parts of the blocks only thin filaments were found. The thin filaments were identified as actin by the formation of arrowhead complexes after incubation with heavy meromyosin.