Biomedical subjects

E Kam

Publications and source records attributed to E Kam.

17 recordsLinked to original sources

PubMed2004

Comparison of Propofol-Lipuro with propofol mixed with lidocaine 10 mg on propofol injection pain.

A common drawback of propofol is pain on injection and lidocaine is commonly mixed with propofol to reduce its incidence and severity. We conducted a randomised, prospective, double-blind study to compare injection pain following the administration of two different formulations of propofol in 200 unpremedicated ASA I-III adult patients scheduled for elective surgery under general anaesthesia. Patients were allocated randomly into two groups to receive either Propofol-Lipuro without added lidocaine or Diprivan mixed with lidocaine 10 mg. Five ml of the study solution was injected at a constant rate over 15 s and patients graded any associated pain or discomfort using a four-point verbal rating scale. The incidence of propofol injection pain was virtually identical in both study groups with 37/98 (38%) patients experiencing pain or discomfort following Propofol-Lipuro compared with 35/98 (36%) after Diprivan (p = 0.88). We observed no significant difference in pain scores between the groups (p = 0.67). Moderate or severe injection pain was experienced by 12/98 (12%) patients given Propofol-Lipuro compared with 8/98 (8%) given Diprivan (p = 0.48).

Adolescent↗

PubMed2001

Survey of patient satisfaction in adult psychiatric outpatient clinics.

A client satisfaction survey was undertaken in two adult psychiatric outpatient clinics. The anonymous self-report questionnaire covering demographic, setting and satisfaction with service variables was endorsed by 203 participants. The mean age of the subjects was 42.5 +/- 19 years, with a small majority (58.6%) of females. Overall satisfaction with psychiatric care was high (79.8%). None of the demographic or setting variables correlated significantly with satisfaction. Psychoeducation was significantly correlated with level of satisfaction with services. These findings further emphasize the importance of psychoeducation by service providers in mental health.

Adult↗

PubMed1998

Neutrophil elastase cleavage of human factor IX generates an activated factor IX-like product devoid of coagulant function.

In preliminary studies, the generation of thrombin in vivo was found to induce a 92% loss of functional activity of factor IX (F.IX) despite the detection by Western blotting of a product resembling activated F.IX (F.IXa) and a 25% increase in F.IX antigen levels (Hoogendoorn et al, Thromb Haemost 69:1127, 1993 [abstr]). These changes were associated with evidence of increased elastase availability. To study the possibility that these two observations were related, a detailed physical and functional characterization of the hydrolysis of purified human F.IX by human neutrophil elastase (HNE) was performed in vitro. An activated partial thromboplastin time (aPTT) clotting assay demonstrated that, although HNE eliminated the potential of F.IX to be activated, it only marginally reduced the F.IXa activity. Reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) indicated that HNE treatment of F.IX generated cleavage products of 30 and 20 kD that could not be distinguished from the respective heavy and light chain peptides that were identified in parallel studies when F.IX was activated by activated bovine F.XI (F.XIa), one of its physiological activators. In addition, nonreducing SDS-PAGE demonstrated that HNE-treated F.IX formed no complexes with antithrombin III (ATIII) in the presence of heparin. Furthermore, HNE-treated F.IX was unable to (1) bind the active site probe p-aminobenzamidine; (2) hydrolyze the synthetic peptide substrate CH3SO2-Leu-Gly-Arg-p-nitroanilide; and (3) activate human factor X (F.X). In contrast to dansyl-Glu-Gly-Arg-chloromethyl ketone (dEGR)-inactivated F.IXa, HNE-treated F.IX (0.01 to 10,000 pmol/L) failed to inhibit the clotting activity of F.IXa (10 pmol/L) in the aPTT. NH2-terminal sequencing indicated that HNE cleaved human F.IX at Thr140, Thr144, Ile164, Thr172, and Val181. The cleavages at Thr140/Thr144 and at Thr172/Val181 are both very close to the normal F.XIa alpha-(Arg145) and beta-(Arg180) cleavage sites, respectively. In summary, the results suggest that the activatability of F.IX is eliminated after cleavage by HNE and that the inability of HNE-treated F.IX to support F.IXa-like coagulant function is a consequence of improper active site formation. These in vitro observations support the possibility that increased HNE cleavage of F.IX in vivo may contribute to the disregulation of hemostasis that occurs in conditions such as disseminated intravascular coagulation (DIC).

Animals↗

PubMed1997

Protein phosphatase activity in human keratinocytes cultured from normal epidermis and epidermis from patients with harlequin ichthyosis.

We investigated serine/threonine protein phosphatase (PP) activity and the expression of PP2A during growth and differentiation of epidermal keratinocytes in culture. Keratinocyte PP activity was strongly inhibited by calyculin A and okadaic acid, indicating that the activity was mainly due to PP2A and PP1. The phosphatase activity decreased to about 20% of the initial (day 1) level by the time of confluence and to about 10% at day 7 postconfluence. In contrast to activity, the level of expression of the PP2A catalytic subunit protein and the mRNA for the two isoforms increased slightly over the period of growth. Keratinocyte differentiation was shown by a significant increase in profilaggrin expression after confluence. Keratinocytes were also cultured from individuals affected with harlequin ichthyosis. This severe hyperkeratotic skin disorder has abnormal lipid structures and is blocked in the PP2A-dependent conversion of phosphorylated profilggrin to the non-phosphorylated filaggrin. The PP activity in harlequin cultures was lower than in normal cultures (about 20% of the subconfluent normal control value) and decreased even further in confluent cultures. In contrast, the level of expression of the PP2A catalytic subunit protein and mRNA for the two isoforms was similar to that of normal keratinocytes and increased with confluence. These results suggest that PP activity in keratinocytes is regulated in a post-translational manner; they also support the possibility of impaired or reduced function of PPs in harlequin ichthyosis.

Blotting, Northern↗

PubMed1993

Harlequin ichthyosis. Variability in expression and hypothesis for disease mechanism.

BACKGROUND: Harlequin ichthyosis is an inherited skin disorder that usually results in death shortly after birth. Although the clinical features of this disorder are well described, the underlying molecular basis is not understood. In this article, we discuss the results of the latest histologic, immunochemical, and Western immunoblotting studies done in our laboratory and propose a hypothesis for molecular basis of this disorder. OBSERVATIONS: Previous experiments done in our laboratory show suggestive evidence for defective lipid synthesis and protein dephosphorylation in harlequin ichthyosis. Our latest study shows that the catalytic subunit of one of the most prevalent protein phosphatase, type 2A protein phosphatase, appears to be altered in some cases of type 2 harlequin ichthyosis. CONCLUSIONS: Based on these observations and the known functions of protein phosphatase in keratinocytes, we hypothesize that the underlying molecular basis of harlequin ichthyosis may be related to mutations affecting protein dephosphorylation. We further describe approaches by which this hypothesis can be tested.

Cells, Cultured↗

PubMed1993

Disposition and orientation of ductin (DCCD-reactive vacuolar H(+)-ATPase subunit) in mammalian membrane complexes.

The disposition and orientation of mouse ductin (the subunit c of the vacuolar H(+)-ATPase) in gap junctions has been examined. Like the Nephrops norvegicus (arthropod) form, mouse ductin in the intact junctional structure is resistant to high levels of nonspecific proteinase, suggesting that it is for the most part buried in the bilayer. Antisera to an octapeptide near the N-terminus cross-react with ductins in gap junction preparations from four different mouse tissues, from chicken and Xenopus laevis liver, and from N. norvegicus hepatopancreas. The antisera and antibodies, affinity purified against the octapeptide, agglutinate isolated gap junctions, suggesting that the N-terminus is located on the exposed surface, equivalent to the cytoplasmic face of an intercellular gap junction. The antibodies also block dye coupling when injected into cells in culture, confirming the cytoplasmic location of the epitope. The lipophylic reagent dicylohexyl carbodiimide (DCCD), which targets carboxyl groups within the membrane and selectively reacts with ductin in N. norvegicus gap junction preparations, rapidly inhibits junctional communication. Bafilomycin A1, which inhibits V-ATPase and stops vacuolar acidification, does not affect dye coupling, showing that the inhibition seen with antibodies and DCCD is not an indirect consequence of their action on the ductin of V-ATPase. Consistent with this interpretation the anti-peptide antibodies do not bind to intact chromaffin granules or inhibit their V-ATPase activity, but do bind to osmotically disrupted granule membrane. This suggests that ductin has an orientation (N-terminus pointing away from the cytoplasm) in the vacuolar membrane opposite to that in the gap junction membrane.

Amino Acid Sequence↗

PubMed1993

Identification of rat epidermal profilaggrin phosphatase as a member of the protein phosphatase 2A family.

The aggregation of cellular intermediate filaments is an important step in the terminal differentiation of keratinocytes. It has been shown that epidermal filaggrin can cause intermediate filaments to aggregate in vitro and may also have the same function in vivo. Filaggrin is derived via dephosphorylation and proteolysis from a highly phosphorylated precursor, profilaggrin, which is found in the granular layer of the epidermis. Using casein kinase II phosphorylated filaggrin as substrate, a profilaggrin phosphatase has been partially purified from rat epidermal homogenate by three chromatographic steps (DE52, hydroxylapatite and S200 gel filtration). Profilaggrin phosphatase activity eluted from the last column has a Km of 0.12 mM and a Vmax of 8 nmol/mg/min with respect to phosphofilaggrin. Results obtained by initial rate analysis showed that the enzymatic activity is not affected by phospho-tyrosyl phosphatase inhibitors and the active fractions preferentially dephosphorylate the alpha subunit of phosphorylase kinase which has been phosphorylated by cAMP-dependent kinase. These results suggest that epidermal profilaggrin phosphatase is not a phospho-tyrosyl phosphatase or a type 1 phospho-seryl/phospho-threonyl phosphatase. Dephosphorylation is not affected by EDTA, calcium or magnesium, but is very sensitive to okadaic acid inhibition (IC50 = 80 pM), suggesting that the enzymatic activity is related to that of the protein phosphatase 2A (PP2A).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Communication compartments in hair follicles and their implication in differentiative control.

Observations on hair follicles presented in this paper show that boundaries to junctional communication are formed between groups of cells following different pathways of differentiation. The patterns of junctional communication in the bulbs of rat vibrissa follicles and human hair follicles were studied by microinjection of the fluorescent tracer dye Lucifer Yellow CH. Dye spread was extensive between undifferentiated cells of the hair bulb matrix but communication boundaries were found between groups of morphologically distinct cells. For example, boundaries to dye spread were observed between undifferentiated matrix cells and cells in the early stage of differentiation into the inner root sheath, between Huxley's and Henle's layers in the early inner root sheath and between cells of the cuticle and cortex of the hair. Dye did not spread between epithelial cells of the hair bulb and mesenchymal cells of the connective tissue sheath or dermal papilla. The patterns of dye spread became more complex (increased boundary formation and subcompartmentation) as differentiation progressed in higher regions of the hair bulb. The observed communication can be related to previous ultrastructural studies by others on the distribution of gap junctions in the wool follicle. These results show that junctional communication, with its consequent intercellular spread of small ions and molecules, is associated with uniformity of expression and behaviour within cell populations and that interruption of communication through the formation of boundaries and communication compartments is temporally and spatially related to the production of subpopulations of cells committed to the expression of different phenotypes.

Animals↗

PubMed1989

Tissue-specific regulation of junctional communication in the skin of mouse fetuses homozygous for the repeated epilation (Er) mutation.

Mouse embryos homozygous for the repeated epilation (Er) gene have abnormally developed skin characterised by hyper-proliferation and incomplete differentiation of the epidermis. In this report, we have studied the patterns of junctional communication in the skin of these mutants to see if the loss of control of proliferation/differentiation is associated with any altered patterns of communication. Using the dye-injection technique we have shown that, compared to normal skin, junctional communication among dermal cells of Er/Er mutants is greatly reduced and the frequency of dermal-epidermal communication is, on the other hand, increased. These results support our previously proposed model, which suggests that selective regulation of junctional communication can be a component of proliferative control in a complex tissue.

Animals↗

PubMed1988

Effects of the tumour promoter 12-O-tetradecanoylphorbol-13-acetate on junctional communication in intact mouse skin: persistence of homologous communication and increase of epidermal-dermal coupling.

12-O-Tetradecanoylphorbol-13-acetate (TPA) has been reported to inhibit junctional communication in some cell cultures. In view of the tumour-promoting activity of TPA in rodent skin, these results have interesting implications for the possible role of junctional communication in tumorigenesis. However, little is known about the effect of TPA on junctional communication in intact skin. Using the technique of iontophoretic injection of Lucifer Yellow CH, we have now studied the patterns of junctional communication in skin treated with TPA. In this paper we report results of such experiments showing that junctional communication in skin persists 4 h after treatment with TPA. In addition, TPA unexpectedly increases communication across the dermal-epidermal boundary. These effects of TPA can still be observed 24 h after treatment. In light of these results, we discuss the possible role of junctional communication in the control of proliferation of normal and pathological tissues.

Animals↗

PubMed1988

Junctional communication and cellular differentiation.

Gap junctions provide pathways of direct cell to cell communication in the tissues of metazoan animals. Cells joined by gap junctions share their small ions and molecules but can maintain distinctive activities through expression of different macromolecules which are too large to pass through the junctions. The junctional channels are made of a tissue invariant, evolutionarily conserved 16-18 k protein but the formation and maintenance of active coupling also requires one or more connexins, a family of tissue-specific proteins ranging in size from 21 k to 70 k. Junctions can be isolated as complexes containing both types of protein by mild procedures using high pH but the connexins can be removed by detergent, urea and protease treatment without destroying the characteristic junctional-morphology of hexagonally packed channels in the double membrane structures. There is also some evidence for the participation in the complex of tissue-specific proteoglycans which perhaps interact with the tissue-specific connexins and account for specificity of junction formation. Such specificity in mixed cultures leads to the production of communication compartments, groups of cells joined by junctions but separated by reduced trans-boundary coupling from cells in adjacent compartments. Compartmentation also occurs in vivo resulting in specific patterns of junctional communication which have been mapped in most detail in mouse skin. These mapping data and the changes which are associated with abnormal proliferation have lead to new ideas on intercellular control.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1987

Molecular structure of the gap junctional channel.

The proteins in various gap junctional preparations from rodent liver have been analysed by two-dimensional peptide mapping and immunoblotting. Only the protein of relative molecular mass (Mr) 16,000 (16K) is found in all gap junctional isolates, and it is unrelated to the 27K protein. The absence of the 27K protein and any of its fragments from trypsin-treated preparations suggests that this protein does not directly contribute to gap junctional structure. Peptide mapping and immunoblotting of the 16K proteins isolated from various tissues and species and of the arthropod 18K protein present in gap junctional preparations from Nephrops norvegicus show that these proteins constitute a family of related junctional proteins. A site-specific antiserum raised against the N-terminal octapeptide of the 16K protein from mouse liver cross-reacts with all 16K and 18K forms of the junctional protein so far tested, suggesting that this particular antigenic determinant is highly conserved. Immuno-localization studies show that the N-terminus is most likely located on the cytoplasmic aspect of the junction and is available to Pronase digestion.

Animals↗

PubMed1987

Patterns of junctional communication in animal tissues.

Gap junctions provide pathways for direct communication between cells in almost all animal tissues. The junctional channels are freely permeable to small ions and molecules but not to macromolecules. A coupled cell population is as a consequence a partial syncytium, within which metabolites, cofactors, small control molecules and inorganic ions can all diffuse freely through the combined cytoplasmic compartment, while intracellular macromolecules remain relatively fixed in space, in the cells where they are synthesized. Little is known about the extent and distribution of these communication compartments in tissues. To further understand their significance, we have mapped the patterns of junctional communication in skin by intracellular injection of the tracer dye Lucifer Yellow. Cells in the dermal layer are widely coupled, with dye spreading through hundreds of cells in a few minutes. Dye spread in the epidermis is much more restricted, passing in the same time into only a few cells. Dermal-epidermal coupling is not normally detected but is occasionally seen in new-born mouse skin when hair follicles are starting to invaginate. Dermal cells are coupled to a band of follicular cells near the base of mature hair follicles, whereas sebaceous glands appear to be coupled units, isolated from surrounding cells.

Animals↗

PubMed1987

Patterns of junctional communication in skin: studies on cultured keratinocytes.

Junctional communication has long been suggested to play a role in coordinating the development of multicellular tissues. A better understanding of the patterns of communication between cells in such tissues is important for the identification of areas where this process may have a role. We have investigated the patterns of communication in cultures of human epidermal keratinocytes by iontophoretic injection of Lucifer Yellow CH, using involucrin expression as a marker of cells undergoing terminal differentiation. Cells that lack involucrin (i.e., the basal, proliferating cells) transfer dye preferentially to other involucrin-negative cells, whereas involucrin-positive cells either are not coupled or transfer dye with similar frequency to involucrin-positive and involucrin-negative neighbors. This decrease in communication associated with terminal differentiation was observed in both the presence and the absence of assembled desmosomes. Our observations lead us to speculate that loss of junctional communication may influence the commitment of basal keratinocytes to terminal differentiation.

Cell Communication↗

PubMed1986

Retinoic acid inhibits junctional communication between animal cells.

Retinoic acid inhibits junctional communication between a variety of vertebrate cell types in culture. It reduces the intercellular transfer of 3H-nucleotides between Syrian hamster kidney fibroblasts (BHK 21/13), Chinese hamster lung fibroblasts (V79), rat liver epithelial cells (BRL), Swiss mouse embryo fibroblasts (3T3), rainbow trout gonadal fibroblasts (RTG2) and Xenopus embryo fibroblasts (Xen). It also reduces metabolic cooperation between hypoxanthine-guanine phosphoribosyl transferase deficient mutant and wild-type BHK cells. The inhibition is rapid (intercellular transfer of iontophoretically injected Lucifer Yellow CH between BRL cells is completely blocked after the cells have been exposed to 10(-4) M retinoic acid for 5 min), and is fully reversed when the drug is removed. Based on these results and the observation that the amount of gap junctional protein isolated from cells grown in the presence of retinoic acid for 1 h is the same and after 24 h is increased (1.3- to 3.1-fold) compared with the amount isolated from untreated cells, we suggest that the inhibitory effect is mediated by the reversible closure of junctional channels.

1-Octanol↗

PubMed1986

Patterns of junctional communication in skin.

The patterns of junctional communication in whole skin have been studied by iontophoretic injection of the fluorescent dye Lucifer Yellow CH into excised strips of tissue from newborn (less than 48-h) mice. Sections of injected specimens embedded in resin show that: coupling in the dermis is extensive, the injected dye spreads to more than 500 cells in 5 min; coupling in the epidermis is more restricted, dye spreads to less than 25 cells in 5 min; cells of the epidermis are not detectably dye-coupled to the cells of the dermis except at occasional restricted sites, probably related to hair follicle development; immature sebaceous gland cells are coupled but appear to be isolated from other cell types; and endothelial cells in the dermis are coupled to surrounding fibroblasts.

Animals↗

PubMed1985

Communication compartments in mixed cell cultures.

Mixed cultures of epithelial (BRL) cells and fibroblasts (BHK), which sort themselves out into separate domains of each cell type, form communication compartments. Electrical coupling, dye coupling and metabolic coupling measurements have been used to show that small ions and molecules can move freely via intercellular junctions between all the cells in a domain, while their movement across the boundaries between domains is severely restricted. Metabolic coupling is the most sensitive method for detecting trans-boundary communication but the results obtained from all three methods are compatible. The data suggest the reduced transfer across the boundaries is due to fewer channels, resulting from a lower frequency of junction formation between heterologous cells, rather than to channels of smaller diameter. Concentration gradients of small cytoplasmic molecules can be established within these communication compartments which are similar to those predicted to explain pattern formation in developing systems. It is suggested that the cell surface features which cause this sorting out are also responsible for the reduced frequency of heterologous junction formation and hence for compartmentalization.

Animals↗