Lesional activation of Tc 17 cells in Behçet disease and psoriasis supports HLA class I-mediated autoimmune responses.

BACKGROUND: Behçet disease (BD) presents with lymphocytic and neutrophilic vasculitis of unknown aetiology. HLA-B*51, the endoplasmic reticulum aminopeptidase 1 (ERAP1), and interleukin 23 receptor (IL23R)/IL12R are genetic risk factors. IL-23 regulates IL-17A, which controls the recruitment and activation of neutrophils. OBJECTIVES: To determine pathological changes in BD skin lesions related to the complex genetic predisposition. METHODS: We characterized the expression of IL-17A and IL-23A in various cell types by immunohistological double staining of sections from papulopustular skin lesions of acute attacks of BD and psoriasis vulgaris lesions, another HLA-class I-associated T-cell-mediated autoimmune disease in which excessive T-cell-derived IL-17A production promotes neutrophil activation. RESULTS: We found that in BD lesions, as in psoriasis, actively expanding CD8+ T cells were the predominant source of IL-17A. IL-17A+ CD8+ T (Tc 17) cells outnumbered infiltrating IL-17A+ CD4+ T cells. Unlike the epidermal localization of CD8+ T cells in psoriasis, Tc 17 cells in BD lesions mainly infiltrated the perivascular tissue and the blood vessel walls of dermis and subcutaneous tissue. They co-localised with a marked IL-23A expression by CD11c+ dendritic cells and CD68+ macrophages. IL-17A expression was associated with extensive recruitment of neutrophils around blood vessels that formed neutrophil extracellular traps (NETs). CONCLUSIONS: In BD, the genetic predisposition may mediate antigen-specific activation and differentiation of a Tc 17 response, possibly targeting endothelial (auto)antigens. Neutrophils recruited by IL-17A in this process may enhance tissue damage by extensive NET formation (NETosis). Thus, the IL-23/IL-17 axis presumably controls neutrophilic inflammation in BD vasculitis in the context of a predominant antigen-specific CD8+ T-cell response.

Proposal of a new classification system for complete mesocolic excison in right-sided colon cancer.

BACKGROUND: Although complete mesocolic excision has been performed for 10 years there remains no published prospective data. The lack of a classification which includes completeness of mesocolic tissue removal as well as plane of surgery contributes to the problem of comparing studies. The aim of the present study was to develop such a classification for right hemicolectomy. METHODS: In a prospective, non-randomized trial we collected specimens of right hemicolectomies from 38 German hospitals between February 2012 and October 2016. The degree of radicality of resection was reported. Photographs were taken of the specimens. After screening the images it became apparent that the specimens could be divided into four main groups according to the degree of missing mesocolic tissue, and three subgroups reflecting the plane of surgery. RESULTS: Of 1373 patients 1097 images were available. Grading was possible in 1077 (98.2%). Distribution was Type 0 (best) 38.6%, Type I 43.3%, Type II 8.5%, Type III (poorest) 7.8%. Surgery was considered to be in a suboptimal plane of surgery in 15.2% overall, highest in Type III (37%) and lowest in Type 0 (7.8%, p < 0.001). CONCLUSIONS: The proposed classification may be a relevant tool for the further investigation of CME for right colon cancer because it allows us to differentiate the aspects of lymphadenectomy and the preservation of the integrity of the mesocolon.

Prevalence of and factors associated with frailty and disability in older adults from China, Ghana, India, Mexico, Russia and South Africa.

BACKGROUND: The severe burden imposed by frailty and disability in old age is a major challenge for healthcare systems in low- and middle-income countries alike. The current study aimed to provide estimates of the prevalence of frailty and disability in older adult populations and to examine their relationship with socioeconomic factors in six countries. METHODS: Focusing on adults aged 50+ years, a frailty index was constructed as the proportion of deficits in 40 variables, and disability was assessed using the World Health Organization Disability Assessment Schedule (WHODAS 2.0), as part of the Study on global AGEing and adult health (SAGE) Wave 1 in China, Ghana, India, Mexico, Russia and South Africa. RESULTS: This study included a total of 34,123 respondents. China had the lowest percentages of older adults with frailty (13.1%) and with disability (69.6%), whereas India had the highest percentages (55.5% and 93.3%, respectively). Both frailty and disability increased with age for all countries, and were more frequent in women, although the sex gap varied across countries. Lower levels of both frailty and disability were observed at higher levels of education and wealth. Both education and income were protective factors for frailty and disability in China, India and Russia, whereas only income was protective in Mexico, and only education in South Africa. CONCLUSIONS: Age-related frailty and disability are increasing concerns for older adult populations in low- and middle-income countries. The results indicate that lower levels of frailty and disability can be achieved for older people, and the study highlights the need for targeted preventive approaches and support programs.

Effect of preorganization on the affinity of synthetic DNA binding motifs for nucleotide ligands.

Triplexes with a gap in the purine strand have been shown to bind adenosine or guanosine derivatives through a combination of Watson-Crick and Hoogsteen base pairing. Rigidifying the binding site should be advantageous for affinity. Here we report that clamps delimiting the binding site have a modest effect on affinity, while bridging the gap of the purine strand can strongly increase affinity for ATP, cAMP, and FAD. The lowest dissociation constants were measured for two-strand triple helical motifs with a propylene bridge or an abasic nucleoside analog, with Kd values as low as 30 nM for cAMP in the latter case. Taken together, our data suggest that improving preorganization through covalent bridges increases the affinity for nucleotide ligands. But, a bulky bridge may also block one of two alternative binding modes for the adenine base. The results may help to design new receptors, switches, or storage motifs for purine-containing ligands.

Deletion of the late cornified envelope genes LCE3B and LCE3C may promote chronic hand eczema with allergic contact dermatitis.

BACKGROUND: Genetically determined defects in epidermal skin barrier function may contribute to the development of irritant and/or allergic contact dermatitis in chronic hand eczema (CHE). OBJECTIVES: To assess whether a deletion in the late cornified envelope genes LCE3B and LCE3C may constitute a genetic predisposition for the development of CHE or any of its subtypes. PATIENTS AND METHODS: A total of 153 German patients with clearly defined CHE subtypes and 268 healthy individuals were screened for the deletion LCE3C_LCE3B-del by allele-specific polymerase chain reaction. RESULTS: Classification of the patients by etiologic subtypes revealed an association between the LCE3C_LCE3B-del allele and CHE due to allergic contact dermatitis. In this subtype, 19/37 patients (51.4%) were homozygous deletion carriers, 11/37 (29.7%) were heterozygous carriers, and just 7/37 (18.9%) were wild-type individuals. Compared to the other CHE subgroups and the healthy control group (homozygous, 88/268 [32.83%]; heterozygous, 133/268 [49.63%]; and wild-type, 47/268 [17.54%]), the prevalence of LCE3C_LCE3B-del in these patients reached statistical significance (P = .03977), as did homozygous deletion carrier status (P = .01044 for other subtypes and P = .02695 for controls). CONCLUSIONS: A deletion of LCE genes may promote the development of allergic contact dermatitis, which is a form of CHE involving delayed-type hypersensitivity.

Filaggrin mutations may confer susceptibility to chronic hand eczema characterized by combined allergic and irritant contact dermatitis.

BACKGROUND: The pathogenesis of chronic hand eczema (CHE) is multifactorial and involves both endogenous predisposition and environmental triggers. OBJECTIVES: Filaggrin is a structural protein of the cornified envelope and important for the formation of the epidermal skin barrier. The aim of this investigation was to evaluate the role of mutations in the filaggrin gene (FLG) in the development of CHE. METHODS: In total, 122 German patients with clearly defined CHE subtypes were screened for the FLG variants R501X and 2282del4 by polymerase chain reaction and restriction enzyme digest analysis. The prevalence of these variants in CHE patients was compared with that in 95 healthy individuals. RESULTS: Overall, allele frequency and the number of mutation carriers were similar in both the CHE and control groups. When classified according to clearly defined CHE subtypes, however, the nonfunctional FLG variants showed an association with CHE involving an aetiological combination of contact allergy and irritant factors [P = 0.04; P (exact test) = 0.06; P (difference in rates) = 0.09; 95% confidence interval (CI) 0-56.8)], or with excessive daily exposure to water and irritants [P = 0.003; P (difference in rates) < 0.001; 95% CI 29.3-67.9]. CONCLUSION: Heterozygosity for nonfunctional mutations in the FLG gene may contribute to the manifestation and maintenance of a particular CHE subtype that is characterized by the combination of allergic and irritant contact dermatitis.

The primary properties?

Science is concerned not just with objects but also with their various aspects, such as their colors, temperatures, sizes, and shapes. These aspects, or properties, are generally thought to be of at least two kinds: primary properties, such as shape, size, and motion and secondary ones, such as temperature and color. However, there is little agreement on just what the difference is between these two kinds of properties. An argument has recently been put forth that assumes the two kinds of properties differ only in that the secondary ones are dependent on the conditions under which they are observed. This paper suggests, however, that since the primary properties depend on conditions, too, any argument based on this assumption is flawed. There are two main traditional accounts of the distinction. The first account of the distinction-the ideas of primary qualities resemble the properties that cause these ideas, whereas those of the secondary ones do not resemble them-relies on what to many is an implausible assumption, that some of our ideas resemble features of the world and we can speak intelligibly about such resemblances. This paper suggests that we cannot, and hence we have no more reason to believe that the ideas of the primary qualities resemble aspects of the world than we do that those of the secondary qualities do so. The second traditional account-primary properties are those a body has, however small it might be, whereas secondary properties are a consequence just of relations between bodies-fares better than the other two accounts. However, the notion of force posed a serious problem for this second basis for the distinction, one that has, astonishingly, hardly found its way into the philosophical literature.

Dominant lesional T cell receptor rearrangements persist in relapsing psoriasis but are absent from nonlesional skin: evidence for a stable antigen-specific pathogenic T cell response in psoriasis vulgaris.

In a previous study we reported that clonally expanded T cell receptor beta-chain rearrangements characterized the T cell receptor usage in skin lesions of psoriasis vulgaris and indicated antigen-specific T cell selection. To assess the relevance of clonal T cell expansion for disease progression, we now determined if select clonal T cell receptor rearrangements persisted over time and were present in nonlesional skin. Sequential biopsies were taken from psoriatic skin lesions of two patients. V-D-J junctional regions of T cell receptor beta-chain variable region gene families 2, 3, 6, 13S1, and BV17 were cloned and sequenced, as these particular BV gene families are preferentially selected in psoriatic skin lesions. The lesional T cell receptor rearrangements were compared with the T cell receptor usage in nonlesional skin and in blood. Several T cell receptor beta-chain rearrangements with high transcript frequency in the first lesional biopsy were again found in sequential lesional biopsies taken as much as 3 y later from psoriasis relapses. Only T cell receptor beta-chain rearrangements with low transcript abundance showed variability in that several clones appeared for the first time or disappeared. Although nonlesional skin also exhibited a restricted T cell receptor usage with clonal T cell receptor rearrangements, the T cell receptor usage in lesional and nonlesional skin differed nearly completely. The select lesional recurrence of identical T cell receptor rearrangements reveals that inflammation in psoriasis involves the same clonally expanded T cell populations and the same antigens over prolonged periods of time. It hereby suggests that specifically recruited and locally expanded T cell clones are permanently involved in psoriatic inflammation and may play a crucial part in disease perpetuation.

The adhesive properties of recombinant soluble L-selectin are modulated by its glycosylation.

The leukocyte adhesion molecule L-selectin, which mediates the initial steps of leukocyte attachment to vascular endothelium, is intensely glycosylated. Different glycoforms of L-selectin are expressed on different leukocyte subsets and differences in L-selectin glycosylation appear to be correlated with the leukocyte's ability to attach to different endothelial targets. In the present study we addressed the question whether glycosylation of L-selectin influences L-selectin-ligand interactions. To obtain different glycoforms of L-selectin, recombinant proteins were expressed both in the baby hamster kidney (BHK) cell line and in the human myelogenous cell line K562, resulting in sL-sel[BHK] or sL-sel[K562], respectively. The glycosylation characteristics of the purified proteins were determined. The most striking differences in glycosylation were seen in the terminal sialylation. Each of the two proteins carried sialic acids in the alpha 2-3 position, while alpha 2-6-bound sialic acids were found exclusively on sL-sel[K562]. To investigate their adhesive properties, both recombinant sL-selectins were used in cell adhesion assays and interactions with the ligands present on various hematopoietic cell lines or activated human cardiac microvascular endothelial cells were examined. The binding capacity of sL-sel[K562] was about 1.6 fold higher compared to sL-sel[BHK] under static as well as under flow conditions. These findings indicate that the terminal sialylation pattern of L-selectin modulates its binding characteristics.

A DEFICIENS homolog from the dioecious tree black cottonwood is expressed in female and male floral meristems of the two-whorled, unisexual flowers.

We isolated PTD, a member of the DEFICIENS (DEF) family of MADS box transcription factors, from the dioecious tree, black cottonwood (Populus trichocarpa). In females, in situ hybridization experiments showed that PTD mRNA was first detectable in cells on the flanks of the inflorescence meristem, before differentiation of individual flowers was visually detectable. In males, the onset of PTD expression was delayed until after individual flower differentiation had begun and floral meristems were developing. Although PTD was initially expressed throughout the inner whorl meristem in female and male flowers, its spatial expression pattern became sex-specific as reproductive primordia began to form. PTD expression was maintained in stamen primordia, but excluded from carpel primordia, as well as vegetative tissues. Although PTD is phylogenetically most closely related to the largely uncharacterized TM6 subfamily of the DEF/APETELA3(AP3)/TM6 group, its spatio-temporal expression patterns are more similar to that of DEF and AP3 than to other members of the TM6 subfamily.

Allometric scaling in small colonies of the scleractinian coral Siderastrea siderea (Ellis and Solander).

Although most physiological traits scale allometrically in unitary organisms, it has been hypothesized that modularity allows for isometric scaling in colonial modular taxa. Isometry would allow increases in size without functional constraints, and is thought to be of central importance to the success of a modular design. Yet, despite its potential importance, scaling in these organisms has received little attention. To determine whether scleractinian corals are free of allometric constraints, we quantified metabolic scaling, measured as aerobic respiration, in small colonies (< or =40 mm in diam.) of the scleractinian Siderastrea siderea. We also quantified the scaling of colony surface area with biomass, since the proposed isometry is contingent upon maintaining a constant ratio of surface area to biomass (or volume) with size. Contrary to the predicted isometry, aerobic respiration scaled allometrically on biomass with a slope (b) of 0.176, and colony surface area scaled allometrically on biomass with a slope of 0.730. These findings indicate that small colonies of S. siderea have disproportionately high metabolic rates and SA:B ratios compared to their larger counterparts. The most probable explanations for the allometric scaling of aerobic respiration are (1) a decline in the SA:B ratio with size such that more surface area is available per unit of biomass for mass transfer in the smallest colonies, and (2) the small size, young age, and disproportionately high growth rates of the corals examined. This allometric scaling also demonstrates that modularity, alone, does not allow small colonies of S. siderea to overcome allometric constraints. Further studies are required to determine whether allometric scaling is characteristic of the full size range of colonies of S. siderea.

Analysis of the TCRBV repertoire of T cells in normal, human skin: evidence for a restricted diversity.

Alpha beta T cells constitute an important component in the first line of immunologic defense in human skin. In order to determine the local selection forces driving T cell diversity, we studied the T cell receptor repertoire in normal human skin and compared it with that of matched blood samples. Using semiquantitative reverse transcription-polymerase chain reaction the expression of T cell receptor beta-chain V genes was determined. The majority of skin, but not blood T cells, revealed a bias towards usage of T cell receptor beta-chain V2 and V6. Whereas sequencing of T cell receptor beta-chain V2 and V6 polymerase chain reaction products showed a heterogeneous clonal distribution within these beta-chain V gene families, the analysis of other selected either over- or underrepresented beta-chain V gene families (BV3, BV12, BV13S1, BV17) revealed numerous identical T cell receptor beta-chain V transcript sequences that were not detected in blood. Restricted T cell receptor diversity in terms of beta-chain V gene preferences or clonal expansion was observed in skin samples of donors from all ages (0.5-87 y). Hence, the repertoire of T cells in normal human skin is apparently subjected to skin-specific selection throughout life. According to our data, this process could involve superantigens, which favor polyclonal accumulation of T cells using certain beta-chain V genes, as well as antigens, which induce clonal T cell expansion. Our results furthermore indicate, that T cell receptor beta-chain V repertoire restrictions do not necessarily result from disease-associated activation of the skin immune system, but could reflect regular mechanisms of immunologic homeostasis within the epithelial surface of the body.

Selection of conserved TCR VDJ rearrangements in chronic psoriatic plaques indicates a common antigen in psoriasis vulgaris.

Psoriasis vulgaris is a common HLA-associated inflammatory skin disease. Although its etiology is still unknown, it is thought to involve T cell-mediated inflammatory mechanisms. In examining the lesional psoriatic TCR beta chain (TCRB) usage in a pair of identical twins concordant for psoriasis, we observed repetitive TCR VDJ rearrangements which indicated antigen-specific oligoclonal T cell expansion. Several of these TCRB rearrangements were identical or highly homologous in the amino acid composition of the complementarity determining region 3 (CDR3), suggesting that T cells with these TCR might be important for disease manifestation. This conclusion was strengthened by TCR analysis of other psoriasis patients. Several repetitive lesional TCRB rearrangements were found that were similar to the conserved CDR3 seen in the twins. Since TCR antigen specificity is largely determined by the beta chain CDR3, selection of T cells with conserved TCRB CDR3 motifs could indicate the presence of a common antigen as a major target of the lesional psoriatic immune response.

Freezing Temperature Effect on Survival of Puccinia graminis subsp. graminicola in Festuca arundinacea and Lolium perenne.

The Willamette Valley in Oregon is a major seed production area for cool-season grasses. Puccinia graminis subsp. graminicola survives over winter on its hosts as uredinial infections and causes epidemics of stem rust, the area's major disease on perennial ryegrass and tall fescue. To determine the possible importance of freezing temperature on rust survival, infected plants taken from the field were subjected to controlled freezing across a range of temperatures representative of those that can occur in the region. After treatment, plants were placed in a warm greenhouse, and the number of actively sporulating pustules was recorded at 3-day intervals for 21 days. The pathogen responded similarly to freezing treatments whether in perennial ryegrass or tall fescue. Compared with the nontreated standard, there was no significant reduction in pustule number after exposure to -3 or -6°C. Exposure of infected plants to -10°C caused a 75 to 90% reduction in rust survival, and exposure to -13°C killed all rust infections in tall fescue and over 99% in perennial ryegrass. The decline in rust survival with temperature was slightly steeper for perennial ryegrass than for tall fescue. A higher absolute number of infections in perennial ryegrass than in tall fescue resulted in higher numbers of surviving infections on perennial ryegrass. Survival of rust infections appeared to be primarily a function of host tissue survival. Between 1960 and 1997, years with winter temperatures as low as -10 or -13°C have occurred in the Willamette Valley with frequencies of approximately 39 and 8%, respectively. We conclude that year-to-year variation in winter temperature could have a significant effect on the survival of the grass stem rust pathogen.

Evidence for an antigen-specific cellular immune response in skin lesions of patients with psoriasis vulgaris.

Psoriasis vulgaris is an inflammatory skin disease characterized by excessively increased keratinocyte proliferation. Several lines of evidence support the idea that T cells infiltrating psoriatic skin lesions play a vital role in the pathogenesis of the disease. To establish whether lesional accumulation and activation of T lymphocytes reflect a specific local immune response, the TCR beta-chain variable (V beta) region gene usage was studied in chronic psoriatic plaques, normal skin, and paired blood lymphocytes. By semiquantitative PCR, we found that overexpression of either or both V beta 2 and V beta 6 gene families characterized the TCR repertoires of normal skin and psoriatic skin lesions. However, sequence analysis of the complementarity-determining region 3 (CDR3) of these V beta gene families demonstrated a marked TCR oligoclonality only in psoriatic lesions, not in normal skin or in blood lymphocytes. The amino acid sequences of the lesional TCR clones revealed that certain conserved junctional motifs were shared by different patients. A second biopsy taken from one of the psoriasis patients 18 mo later from a different anatomical site disclosed that the same TCR clones were again dominating. These data suggest that lesional psoriatic T lymphocytes expressing the prevailing TCR V beta genes represent an oligoclonal T cell subset that expanded from a few progenitor T cells in response to Ag in the skin of psoriasis patients. They are derived from a polyclonal T cell population that, by the expression of V beta 2 or V beta 6 TCR, appears to be predisposed for homing to the skin.

Mitochondrial DNA products among RAPD profiles are frequent and strongly differentiated between races of Douglas-fir.

Racial differentiation and genetic variability were studied between and within the coastal, north interior, and south interior races of Douglas-fir using RAPD and allozyme markers. Nearly half of all RAPD bands scored (13: 45%) were found to be amplified from mitochondrial DNA. They exhibited maternal inheritance among hybrids and backcrosses between the races, and were much more highly differentiated (GST = 0.62 for haplotype frequencies) than were allozymes (GST = 0.26). No evidence of hybridization or introgression was detected where the coastal and interior races come into proximity in central Oregon.

T lymphocytes derived from skin lesions of patients with psoriasis vulgaris express a novel cytokine pattern that is distinct from that of T helper type 1 and T helper type 2 cells.

In various immunological disorders the pathomechanisms of tissue damage are causally associated with specific patterns of locally produced cytokines. To study the molecular and cellular mechanisms involved in the manifestation of psoriasis vulgaris we have assessed the cytokine mRNA profile expressed in lesional psoriatic skin and in T cell clones (TCC) that were established from skin lesions of patients with psoriasis. As demonstrated by use of the polymerase chain reaction (PCR), psoriasis lesions consistently exhibit transcription of a complex pattern of cytokines. It includes mediators selectively produced by T lymphocytes [interferon (IFN)-gamma, tumor necrosis factor (TNF)-beta, interleukin (IL)-2, IL-3 and IL-5] as well as cytokines secreted by various cell types [transforming growth factor (TGF)-alpha/-beta, TNF-alpha, IL-6/-8 and granulocyte-macrophage-colony stimulating factor], while IL-4 is missing. With the exception of TGF-alpha, this cytokine profile was also observed in lesional psoriatic T cell clones yielding supernatants mitogenic for keratinocytes in vitro (MTCC), but not in T cell clones yielding supernatants that inhibited keratinocyte proliferation (STCC). The congruent cytokine expression of psoriatic skin lesions and MTCC emphasizes that inflammation in psoriasis is driven by a sofar unrecognized regulatory T cell subset that may serve to control epidermal regeneration and convey immunosurveillance over epithelial surfaces. It is characterized by the combined expression of IFN-gamma, TGF-beta, IL-2 and IL-5 in the absence of IL-4 and by its selective capacity to enhance keratinocyte proliferation. This newly defined combination of regulatory properties of a distinct T cell population cannot be reconciled with an immune response of the T helper cells (TH)0, TH1 or TH2 type.

Guanosine 5'-O-[S-(4-bromo-2,3-dioxobutyl)]thiophosphate and adenosine 5'-O-[S-(4-bromo-2,3-dioxobutyl)]thiophosphate. New nucleotide affinity labels which react with rabbit muscle pyruvate kinase.

Three new reactive nucleotide analogues with bromo-keto substituents adjacent to a thiophosphate have been synthesized. Guanosine 5'-O-[S-(4-bromo-2,3-dioxobutyl)]thiophosphate (GMPS-BDB), reacts covalently with rabbit muscle pyruvate kinase with complete inactivation and incorporation of 1.8 mol of reagent/mol of enzyme subunit. By contrast, the mono-keto compound, guanosine 5'-O-[S-(3-bromo-2-oxopropyl)]thiophosphate (GMPS-BOP), causes no loss of pyruvate kinase activity. When the analogous adenosyl nucleotide derivatives are incubated with pyruvate kinase, the di-keto compound, adenosine 5'-O-[S-(4-bromo-2,3-dioxobutyl)]thiophosphate (AMPS-BDB), rapidly effects inactivation, whereas the mono-keto compound, adenosine 5'-O-[S-(3-bromo-2-oxopropyl)]thiophosphate (AMPS-BOP), causes no loss of activity. Complete protection against inactivation by GMPS-BDB is provided by phosphoenolpyruvate in the presence of K+ and Mn2+ and the amount of reagent incorporated (0.9 mol/reagent/mol subunit) is reduced to half that observed in the absence of protectants. Gas-phase sequencing of the tryptic peptides purified from inactive GMPS-BDB or AMPS-BDB-modified enzyme gave the cysteine-labeled peptides: C151DENILWLDYK161, and N162IC164K165 as the two major peptide products, with a smaller amount of N43TGIIC48TIGPASR55. Reaction in the presence of the protectants PEP, K+, and Mn2+ yielded Cys164 as the only labeled residue, indicating that inactivation is primarily due to modification of Cys151. We propose that GMPS-BDB (or AMPS-BDB), which may exist in enolized form in aqueous solution, functions as a reactive analogue of phosphoenolpyruvate and GDP (ADP) to target Cys151 in the active site of pyruvate kinase.

The activity of S-thiomethyl modified creatine kinase is due to the regeneration of free thiol at the active site.

Creatine kinase modified by S-methyl methanethiosulfonate and devoid of reactive thiol group has been reported to retain about 18-40% of the activity of the native enzyme. It has now been found that during the reaction catalyzed by the modified enzyme the rate increases with time and if the reaction is allowed to continue sufficiently long, the enzyme eventually recovers full activity. The presence of substrates is not required for the reactivation as suitable dilution after removal of MMTS in excess leads to complete reactivation of the MMTS modified enzyme with the simultaneous regeneration of reactive thiol per each dimeric molecule as shown by determinations with DTNB and IAN. The addition of MMTS during the course of reactivation again inactivates the reactivated enzyme. The activity recovery is therefore due to the regeneration of reactive thiol and it appears that the active-site thiols are essential for the activity of rabbit muscle creatine kinase.

T cell clones from psoriasis skin lesions can promote keratinocyte proliferation in vitro via secreted products.

Psoriasis vulgaris has been recognized lately as an immunologically mediated inflammatory skin disease. To analyze the pathogenetic role of T lymphocytes in the generation of psoriatic skin lesions, 105 T cell clones (TCC) and 10 T cell lines (TCL) were differentially isolated from dermis and epidermis of psoriatic skin specimens. Supernatants prepared from these T cells were studied for their effects on keratinocyte proliferation in vitro. Conditioned media from 14 of 77 epidermal TCC, 7 of which were CD8+, and from 8 of 28 dermal TCC, 5 of which were CD8+, reproducibly enhanced keratinocyte proliferation, with more pronounced mitogenic activities found in dermal TCC. Another 9 epidermal and 3 dermal TCC did not affect keratinocyte growth and supernatants from the remaining clones, as well as from the 5 epidermal and 5 dermal TCL, inhibited keratinocyte replication to varying degrees. Both mitogenic and suppressive activities were largely abolished by addition of an antiserum to interferon-gamma (IFN-gamma), while addition of epidermal growth factor or irradiated psoriatic TCL had little effect on the activities of the supernatants. These studies reveal that a subpopulation of lesional psoriatic T lymphocytes is capable of enhancing keratinocyte proliferation in vitro via secreted products. Their mitogenic capacity most likely requires IFN-gamma, but the ultimate effect is apparently determined by the presence of additional cytokines. Activation of T cells secreting such combinations of factors in vivo may contribute to the keratinocyte alterations characteristic of psoriatic skin lesions.