Skin Window Research Articles

Efficacy of Ascorbic Acid in Chediak-Higashi Syndrome (CHS): Studies in Humans and Mice

The efficacy of ascorbic acid therapy in Chediak-Higashi syndrome (CHS) was assessed in two humans and in the mouse model of this disease. Ascorbic acid (6 g/day) was given for 8 mo to two brothers (26 and 27 yr old) with CHS. The therapy had no effect on the clinical course of these patients and no change in their complete blood count or abnormal leukocyte morphology (giant lysosomes). In addition, the leukocyte dysfunction seen in this disease was not improved by ascorbic acid; skin window responses, chemotaxis, and bactericidal activity remained abnormal and there was no change in lymphocyte function. No abnormality of leukocyte cAMP or cGMP was seen in our patients, and no changes in basal cyclic nucleotides occurred during ascorbic acid therapy. In other studies high doses of intraperitoneal ascorbic acid (20 mg/mouse) prolonged the survival of CHS mice from lethal infection with Candida albicans, although the survival of CHS treated mice was not as good as that of saline-treated controls. Ascorbic acid also improved polymorphonuclear leukocyte chemotaxis and bactericidal activity for Staphylococcus aureus. Despite the improved function there was no change in the morphology of CHS mouse leukocytes after ascorbic acid therapy. In addition, we found no abnormality of basal leukocyte cyclic nucleotide levels and no change in basal cyclic nucleotides after ascorbic acid therapy. These data indicate that although ascorbic acid may have a beneficial effect in mice with CHS, patients with CHS may not respond in a similar manner.

SEM of exudative leucocytes obtained by the skin window technique

SEM of exudative leucocytes obtained by the skin window technique

Defective Neutrophil Migration in Granuloma Annulare, Necrobiosis Lipoidica, and Sarcoidosis

The neutrophil mobility in patients with granuloma annulare, necrobiosis lipoidica, and sarcoidosis was tested using both in vivo and in vitro techniques. Use of a skin window chamber for measurement of chemotaxis in vivo demonstrated defective neutrophil migration in each group. This contrasted with the finding that, in vitro, chemotaxis toward casein and endotoxin-activated sera was normal. The importance of this dissociation is discussed. Abnormal neutrophil accumulation at sites of inflammation may be of importance in the pathogenesis of these granulomatous disorders.

Experimental Studies on Phototoxic and Photoallergic Reaction by the Skin Window Technique

The differentiation of photoallergic contact dermatitis by 3,3', 4', 5-tetra-chlorosalicylanilide from phototoxic contact dermatitis by 8-MOP in guinea pigs was studied by skin window technique. Based upon the results, it may be concluded that the photoallergic reaction can be differentiated from phototoxic reaction by the appearing pattern of basophilic leukocytes.

Detection of cellular immunity derangements in chronic gastritis by a skin window test

An autologous "skin window test," used lately for the study of cellular immunity in cancer, was applied here successfully to 54 patients with upper-gastrointestinal ailments, 48 of whom had a coexistent fundic and/or antral chronic gastritis of varying severity. The diagnoses of gastritis were made by multiple fiber-gastroscopic biopsies. The ether-alcohol-fixed cryostat sections of fundic and antral biopsies were mounted on cover slides and placed on small cutaneous abrasions of the forearm of the same patients for 24--28 hr. The exudates on cover slides and on imprints of the abrasions were read blindly for the mononuclear cell response according to criteria set for this test by Black and Leis (10). A positive reaction was obtained in 8 of the 54 patients using autologous fundic mucosal biopsy. An autologous antral mucosal biopsy gave positive reaction in only 2 of the 26 of the patients in whom it was used. The positive yield of this autologous skin window test in chronic advanced fundic gastritis was somewhat higher than that obtained by other authors using lymphocytes blast transformation or macrophages migration inhibition test in vitro. It was much higher than the yield obtained by others who used skin tests in vivo, with homologous or heterologous gastric mucosal extracts as antigens. The autologous skin window is safe in regard to possible transmission of hepatitis. Its applicability for detection of cellular immunity derangement in chronic gastritis carries promise.

Darier's Disease

Darier's disease is frequently complicated by bacterial skin infections and occasionally by Kaposi's varicelliform eruption. Postulating that defective host immunologic competence might explain these infections, humoral and cell-mediated immunity (CMI) were evaluated in four patients. Humoral immunity was normal as demonstrated by quantitative immunoglobulins, isohemagglutinins, direct skin immunofluorescence, and B-cell counts. The CMI was evaluated by standard delayed type hypersensitivity skin tests, T-cell counts, lymphocyte transformation assays, macrophage inhibition factor (MIF) assays, and skin windows. Blunted lymphocyte blastogenesis, MIF, and skin window response indicated depressed CMI. Polymorphonuclear leukocyte chemotaxis and phagocytosis were normal.

Polymorphonuclear cell function in the various polar types of leprosy and erythema nodosum leprosum.

Polymorphonuclear leukocyte motility, both in vivo and in vitro, and reduction of Nitro Blue Tetrazolium was studied in tuberculoid and lepromatous leprosy patients and a group of lepromatous patients with erythema nodosum leprosum (ENL). A profound defect in random migration, chemotaxis, and chemokinesis was found in lepromatous patients with and without complicating ENL, and marked depletion of skin window migration confirmed these in vitro findings. Tuberculoid patients exhibited a mild defect in polymorphonuclear leukocyte motility. Serum inhibitors of normal polymorphonuclear leukocyte chemotaxis were found in all types of leprosy, but sera from lepromatous and ENL patients were most inhibitory. Resting levels of Nitro Blue Tetrazolium reduction were normal in all three groups. Reconstitution of polymorphonuclear leukocyte cells from normal and ENL patients with ENL serum, however, showed increased Nitro Blue Tetrazolium reduction well above the normal range, whereas reconstitution with normal, lepromatous, and tuberculoid sera failed to increase Nitro Blue Tetrazolium reduction above the normal values.

Angioimmunoblastic lymphadenopathy.A T-cell deficiency

The case of a 49-year-old man with the diagnosis of angioimmunoblastic lymphadenopathy is reported. The patient survived a stormy clinical course. The corticosteroids improved dramatically the clinical picture although the patient developed a staphylococcal septicemia. Before treatment immunological studies were done including quantitation of B and T-cells, antigen stimulation of lymphocytes in vitro, skin tests and skin window. Impairment of cell mediated immunity, decreased T-lymphocytes and increased B-lymphocytes were found. A decreased migration of lymphocytes in the skin window was also found compatible with immunosuppression. A possible presumptive pathogenetic mechanism is described although the cause of this recently described entity remains unknown.

The direct demonstration of histamine release in allergic reactions in the skin using a skin chamber technique

We have adapted a skin chamber technique to permit sampling of fluid at the skin window sites of pollen antigen-induced allergic reactions. Low background levels of histamines are formed in control chambers, whereas significantly increased (p < 0.01) amounts are found within 30 min following ragweed application in sensitized subjects.

Chemotactic Response to Human C3a and C5a Anaphylatoxins

Abstract Peptides C3a and C5a, also known as anaphylatoxins, are derived from C components C3 and C5 and have previously been reported to exhibit chemotactic activity. Virtually no data exist, however, regarding the relative potency of C3a or C5a for stimulating directed cellular movement. The chemotactic response of polymorphonuclear leukocytes (PMN) toward purified human C3a and C5a was determined under both in vitro and simulated in vivo conditions. Leukotactic activity of human C5a and C3a was examined with a modified Boyden chamber assay over a broad concentration range. Human C5a exhibited activity only over the range of 0.04 to 1.7 × 10-8 M. Greater or lesser concentrations of C5a were incapable of stimulating cellular migration. Since C5a is rapidly converted to C5ades Arg by a carboxypeptidase in serum, we also examined this modified anaphylatoxin molecule. For this study C5aArg was formed by digesting C5a with pancreatic carboxypeptidase B (EC 3.4.2.2). Human C5ades Arg is a form of C5a that is inactive as an anaphylatoxin. C5ades Arg was active as a chemotactic factor in the in vitro assay, but only when nonactivated serum was added. Approximately 10 times more C5ades Arg than C5a is required to stimulate a leukotactic response and the magnitude of the maximum cellular response is only about 50% of that produced by intact C5a. It was observed that serum provides a definite helper role for C5ades Arg stimulated chemotaxis under the in vitro assay procedures employed. Human C3a was inactive over a concentration range from 8 × 10-10 to 8 × 10-6 M and trypsintreated human C3 was also active over a range of 4 × 10-8 to 2.4 × 10-6 M. Earlier reports that 1- to 5-min trypsin digestion of C3 generates a chemotactic activity were not confirmed by our studies with human PMN; however, previous observations of activity may have resulted from minor contamination of the C3 with C5. Human C5a was maximally active over a narrow concentration range of 2 to 6 × 10-7 M as estimated in rabbits by utilizing a skin window assay technique (Otani and Hugli, 1977). Human C5ades Arg also stimulated neutrophil accumulation under simulated in vivo conditions but at a slightly higher concentration range of 0.2 to 1.2 × 10-6 M. The C5ades Arg did not require added serum to produce an apparent leukotactic response in vivo, but, as in vitro, activity was not observed at levels above or below the effective concentration ranges indicated for C5a and C5ades Arg C3a was inactive over a concentration range of 5.5 × 10-9 to 5.5 × 10-6 M as an attractant for rabbit neutrophils in the skin window assay. The concentrations of C5a or C5ades Arg that are required to induce chemotaxis under simulated in vivo conditions correspond to the potential serum levels of these factors (e.g., 4 to 5 × 10-7 M). It is suggested from our quantitative measurements that the predominant chemotactic activity generated in human serum during C activation is associated with the C5ades Arg molecule. Therefore, C5ades Arg exhibits the potential for playing a predominant functional role under physiologic conditions as a major humoral chemotactic factor.

Abnormal neutrophil behaviour in sarcoidosis: studies using a quantitative skin window technique

Journal Article Abnormal neutrophil behaviour in sarcoidosis: studies using a quantitative skin window technique Get access R.W. Gange, R.W. Gange St John's Hospital for Diseases of the Skin, London Search for other works by this author on: Oxford Academic Google Scholar M.M. Black, M.M. Black St John's Hospital for Diseases of the Skin, London Search for other works by this author on: Oxford Academic Google Scholar R. McKerron R. McKerron St John's Hospital for Diseases of the Skin, London Search for other works by this author on: Oxford Academic Google Scholar British Journal of Dermatology, Volume 97, Issue s15, 1 December 1977, Pages 15–16, https://doi.org/10.1111/j.1365-2133.1977.tb14296.x Published: 01 December 1977

Evaluation of Congenital Neutropenic Disorders by in Vitro Bone Marrow Culture

The congenital neutropenias are a heterogenous group of diseases whose etiology and pathogenesis are largely unknown. We studied nine neutropenic patients from seven families. Evaluation included peripheral blood cell and differential cell counts, epinephrine and typhoid vaccine stimulation studies. Rebuck skin windows, and bone marrow aspirations for morphological assessment and for in vitro culture in liquid suspension and in agar plates. Parallel cultures were set up with and without colony-stimulating activity (CSA), and peripheral leukocytes were assayed for cellular production of CSA. Patients were initially classified on the basis of their clinical course: benign, mild, moderately severe, or severe disease. One patient in the moderately severe group had an immunoglobulin disorder. Morphologically normal mature granulocytes were seen in bone marrow aspirates of two patients, and maturational defects of varying degree were seen in the remaining seven. Colony formation in agar was markedly reduced below normal in three of seven, moderately reduced in two of seven, and greater than normal in two patients. Colonies in six of seven patients consisted exclusively of macrophages. Marrow from all but one of the nine patients demonstrated poor neutrophil development in suspension culture, and addition of CSA did not result in augmented granulocytic proliferation or maturation. A scheme of normal neutrophil maturation is proposed, and the nine patients were categorized according to this scheme. Four patterns of congenital neutropenia emerged: type 1 was the most benign form of disease with essentially normal clinical and in vitro parameters, and a defect considered to be due to a small committed stem cell pool, abnormal release, or excessive utilization peripherally; type 2 had mild disease with presumed defective committed stem cell differentiation along the granulocyte line; type 3 included benign to severe clinical expression with an apparent defect at the level of the committed granulocyte precursor more severe than in type 2; type 4 disease had varied clinical expression but evidence for a defect at the level of the pluripotent stem cell.

Levels of Carbenicillin, Ticarcillin, Cephalothin, Cefazolin, Cefamandole, Gentamicin, Tobramycin, and Amikacin in Human Serum and Interstitial Fluid

The ability of eight antibiotics (carbenicillin, ticarcillin, cefamandole, cephalothin, cefazolin, gentamicin, tobramycin, and amikacin) to enter human interstitial fluid was evaluated by the skin window technique. All of the antibiotics tested, except cefazolin, which has the highest percentage of protein binding, diffused into the interstitial fluid quite well. This study confirms our previous observation on the effect of high-percentage protein binding on diffusion of a drug into the minimally inflamed extravascular space.

CONGENITAL LEUKOCYTE MOVEMENT DISORDER AND RECURRENT INFECTIONS

A 4½ month old black female with omphalitis at 4 days of age, recurrent infections of the skin, an acquired rectovaginal fistula, marked lymphadenopathy proximal to sites of infection and delayed wound healing had a peripheral WBC count of 70-100,000/mm3 with 70% PMN's and 10% monocytes. PMN's were absent in exudates of infected areas and a Rebuck skin window at 4, 8 and 12 hours. A chemotactic assay under agarose revealed absent directed movement and decreased non-directed movement of PMN's and monocytes. Zymosan activated serum from both patient and normal controls attracted normal cells to the same degree. No serum, plasma or cellular inhibitor to normal PMN movement was found. The patient's mean PMN velocity by direct observation was 2.4 μ/min. (control 11.9 μ/mtn.) and her mean monocyte velocity was 2.1 μ/min. (control 4.5 μ/min.). By light microscopy the patient's cells formed multiple, small lame 11ipodia without developing a single dominant lamel1ipodium; electron microscopy of the cells was normal. Associated findings included marked panhyperimmunoglobulinemia with an IgE of 2600-4300 ng/ml. Assays of the phagocytes' metabolic, phagocytic and bactericidal functions, T and B cell functions and complement system were normal. This black child with severe infections from birth, a markedly elevated IgE without allergic symptoms and defective movement of both PMN's and monocytes represents a congenital and possibly genetically determined abnormality of cell mobility.

Functional evaluation of circulating leukemic cells in acute non-lymphocytic leukemia

Peripheral blood leukemic cells from 32 patients with acute non-lymphocytic eukemia (ANLL) were tested in vitro for their ability to adhere to nylon fibers and to phagocytize latex particles and S. aureus. In vivo migration to skin windows was also assessed. Using heparinized whole blood, leukemic cells from 47% of patients ( 9 19 ) adhered to fibers although the percent adherence was decreased compared to the per cent of granulocyte adherence in these patients. Phagocytosis by leukemic cells of S. aureus and latex was seen in 44% ( 12 77 ) of patients studied and migration to skin windows occurred in 44% ( 10 23 ). Only cells from patients with some degree of monocytic differentia-(monocytic or myelomonocytic leukemia) were functional while blasts from patients with myelocytic, promyelocytic or poorly differentiated leukemia were non-functional. Prominent extramedullarly leukemic involvement was more common in patients with functional monoblasts and on occasion dominated the patient's clinical course. These data indicate that some degree of monocytic functional maturation takes place in many patients with ANLL and may account for the increase in extramedullary infiltration seen in patients with monocytic leukemia. The simple functional tests described may supplement morphologic criteria in the characterization of ANLL.

Basophils in tuberculin and "Jones-Mote" delayed reactions of humans.

Jones-Mote reactions are delayed, erythematous, and mildly indurated cutaneous reactions originally described in humans sensitized by skin injection of heterologous proteins. Similar reactions in guinea pigs contain many basophils and are called cutaneous basophil hypersensitivity. In contrast, guinea pigs immunized with mycobacterial adjuvants have classical tuberculin-type delayed hypersensitivity reactions, which contain few basophils. This has led to a new classification of delayed responses, based largely on the presence or absence of basophils. We induced sensitization for Jones-Mote reactions in 20 normal humans by intradermal injections of keyhole limpet hemocyanin. Skin tests with KLH 1 wk later showed erythematous and indurated delyaed reactions in all subjects. Rebuck skin windows showed specific accumulations of basophils with a delayed time-course in 18 of 20 subjects. In 12 normals sensitized with oxazolone-keyhole limpet hemocyanin conjugates, skin reactions and in vitro lymphocyte stimulation showed carrier and not hapten specificity, suggesting that cutaneous responses were probably mediated by T cells. A comparative study of strongly positive PPD skin tests in patients with tuberculosis showed significant basophil accumulations in five of nine subjects. Thus, basophils occurred in human tuberculin and Jones-Mote reactions and were not a distinguishing feature of Jones-Mote reactions. We suggest that the occurrence of basophils at delayed reactions is under complex regulation and that basophil accumulations are an aspect of delayed hypersensitivity, rather than an indication of a distinctive and separate response.

Defective Polymorphonuclear Leukocyte Chemotaxis and Bactericidal Capacity in a Boy With Recurrent Pyogenic Infections

A 13-year-old boy with a history of recurrent pyogenic infections had abnormalities of polymorphonuclear leukocyte (PMN) function which probably accounted for his susceptibility to infection. PMN phagocytosis and nitroblue tetrazolium dye reduction were normal but glucose 14C oxidation was abnormally increased in resting cells. The patient's PMNs possessed decreased bactericidal activity against Staphylococcus aureus and Escherichia coli. Also documented were decreased PMN chemotactic activity and leukocyte accumulation in Rebuck skin windows. PMN random motility, PMN bone marrow reserve, PMN kinetics, lymphocyte blastogenesis, and delayed cutaneous hypersensitivity were normal. This patients represents another of the growing number of distinct granulocytopathies currently being recognized which may result in increased susceptibility to infection.

Application of scanning electron microscopy to the study of skin window preparations.

Application of scanning electron microscopy to the study of skin window preparations.

NEUTROPHIL DYSFUNCTION IN CROHN'S DISEASE

The migration of neutrophils into skin windows has been examined in patients with Crohn's disease and rheumatoid arthritis and compared with normal. Leucocyte migration was significantly lower in patients with Crohn's disease than in others. This abnormality of neutrophil function in Crohn's disease appears to be secondary to a defective acute inflammatory response as the neutrophils themselves were found to behave normally on in-vitro testing. A weak acute inflammatory response to particulate or antigenic material in the bowel wall could result in the chronic inflammation observed in this condition.

Defective neutrophil motility in children with measles

The random migration and chemotactic ability of neutrophils from ten patients with uncomplicated measles was found to be grossly impaired when compared to a group of normal children. Chemotaxis to endotoxin activated serum and to hydrolysed casein was markedly depressed, but serum from measles patients, when activated by endotoxin, generated and chemotactic activity and did not contain leukotactic inhibitors. The defect in neutrophil motility was confirmed in vivo by abnormal Rebuck skin windows. The defect was temporary, and recovery of normal chemotaxis was observed by the eleventh day after the onset of the rash.