The effects of Unguentum Lymphaticum on skin in patients with obstructive lymphedema of the lower extremities.

Obstructive lymphedema of extremities in humans is characterized by swelling of tissues with lymph stasis and inflammatory infiltrates in skin and subcutaneous tissues. Treatment of the inflammatory component requires application of antiinflammatory drugs. We studied the effect of topical application of Unguentum Lymphaticum (UL) containing antiphlogistic compounds (digitalis, calendulin, hyoscyamine, colchicine and podophyllin) on lymphedematous skin in 33 patients with stage II postinflammatory obstructive lymphedema. A three-week treatment of swollen legs with UL brought about stimulation of epidermal cells with proliferation of keratinocytes, increased numbers of Langerhans cells, accumulation of macrophages in the dermis and activation of infiltrating cells and endothelia. Besides some foci of acanthosis, no degenerative changes were observed in the skin in patients treated for 12 weeks and no changes were observed in the placebo treated groups. Immunohistochemical evaluation of biopsy specimens of inguinal lymph nodes in patients treated for 12 weeks revealed reactive granulocyte and macrophage accumulation in the cortical and paracortical areas. Components of UL inhibited stimulation of blood mononuclear cells in in vitro cultures. UL did not change the spectrum of calf skin bacterial populations. The nonspecific stimulation of skin-associated lymphoid tissue and putative elimination of factors responsible for lymphe-dematous skin inflammation appears to be responsible for the beneficial clinical effect of UL on legs with lymph stasis.

Bacteriological studies of blood, tissue fluid, lymph and lymph nodes in patients with acute dermatolymphangioadenitis (DLA) in course of 'filarial' lymphedema.

Filarial lymphedema is complicated by frequent episodes of dermatolymphangioadenitis (DLA). Severe systemic symptoms during attacks of DLA resemble those of septicemia. The question we asked was whether bacterial isolates can be found in the peripheral blood of patients during the episodes of DLA. Out of 100 patients referred to us with 'filarial' lymphedema 14 displayed acute and five subacute symptoms of DLA. All were on admission blood microfilariae negative but had a positive test in the past. Blood bacterial isolates were found in nine cases, four acute (21%) and five subacute (26%). In 10 acute cases blood cultures were found negative. Six blood isolates belonged to Bacilli, four to Cocci and one was Sarcina. To identify the sites of origin of bacterial dissemination, swabs taken from the calf skin biopsy wounds and tissue fluid, lymph and lymph node specimens were cultured. Swabs from the calf skin biopsy wound contained isolates in nine (47%) cases. They were Bacilli in nine, Cocci in three, Acinetobacter and Erwinia in two cases. Tissue fluid was collected from 10 patients and contained Bacilli in four (40%) and Staphylococci in three (30%). Lymph was drained in four patients and contained isolates in all samples (100%). They were Staphylococcus epidermis, xylosus and aureus, Acinetobacter, Bacillus subtilis and Sarcina. Three lymph nodes were biopsied and contained Staphylococcus chromogenes, xylosus, Enterococcus and Bacillus cereus. In six cases the same phenotypically defined species of bacteria were found in blood and limb tissues or fluids. In the 'control' group of patients with lymphedema without acute or subacute changes all blood cultures were negative. Interestingly, swabs from biopsy wound of these patients contained isolates in 80%, tissue fluid in 68%, lymph in 70% and lymph nodes in 58% of cases. In healthy controls, tissue fluid did not contain bacteria, and lymph isolates were found only in 12% of cases. This study demonstrates that patients with acute episodes of DLA reveal bacteremia in a high percentage of cases. Diversity of blood and tissue bacterial isolates in these patients points to a breakdown of the skin immune barrier in lymphedema and subsequently indiscriminate bacterial colonization of deep tissues and spread to an blood circulation.

Bacteriologic studies of skin, tissue fluid, lymph, and lymph nodes in patients with filarial lymphedema.

Filarial lymphedema is complicated by frequent episodes of dermatolymphangioadenitis (DLA). It is not certain whether DLA is of filarial or bacterial etiology. The frequency of episodic DLA does not depend on the presence or absence of microfilariae. Antibiotic therapy is effective in prevention and treatment of DLA. These observations point to the bacterial rather than filarial etiology of DLA. Skin and lymph node biopsies, tissue fluid, lymph, and blood from patients with chronic filarial lymphedema, and during acute episodes of DLA, were cultured for detection of bacteria. A high prevalence of bacterial isolates from the tissue fluid (64%), lymph (75%), and inguinal lymph nodes (66%) of limbs with filarial lymphedema was found. Bacillus cereus, Staphylococcus epidermidis, S. hominis, S. capitis, S. xylosus, and Micrococcus spp. were the most common isolates. Bacteria were also isolated from the blood of patients with recent episodes of DLA, with strains of the same phenotype and antibiotic sensitivity in all specimens from patients with DLA. Bacterial strains of the same phenotype and antibiotic sensitivity were documented on the toe web surface and in tissue fluid (25%), lymph (26%), or lymph nodes (41%). Increasing prevalence of bacterial isolates in tissue fluid, lymph, and lymph nodes was observed in advanced stages of lymphedema. Bacilli and cocci were sensitive to gentamicin, tetracyline, rifampicin, vancomycin, kanamycin and cotrimoxazole, and least sensitive to penicillin. Blood cultures of patients in the periods between DLA attacks were negative. In healthy controls without edema and episodes of DLA, tissue fluid did not contain bacteria. In lymph, only single colonies of Micrococcus and Acinetobacter were cultured in 12% of the cases. Impaired lymph drainage and lack of elimination of penetrating bacteria may be responsible for progression of lymphedema and recurrent attacks of DLA.

Normal human immune peritoneal cells: subpopulations and functional characteristics.

Normal human peritoneal cells collected during elective laparatomy from patients with gallbladder stones without clinically detectable inflammatory changes were characterized phenotypically with immunocytochemical method and flow cytometry, with special attention paid to the presence of memory cells. The responsiveness of normal PCs to mitogen and, specifically, the role of peritoneal macrophages in this process was studied. The peritoneal cells consisted of 45% of monocytes/ macrophages (CD68+), as many as CD2+ T lymphocytes, 8% CD57+ NK and K 2% CD22+ B, cells. The CD4/CD8 ratio was 0.4. The peritoneal cells did not express interleukin-2 (CD25+) and transferrin receptors (CD71+) on their surface. Approximately 49% of the peritoneal cells were class II MHC antigen positive cells. Two per cent of S100+ dendritic cells were found. Flow cytometric two-colour analysis revealed that the majority of peritoneal CD4+ (92.4%) and CD8+ (73.1%) lymphocytes, while only 50.2% of CD4+ and 30.1% CD8+ peripheral blood cells expressed simultaneously the CD45R0 (UCHL1) molecule, which is characteristic to the memory/effector T-cell subpopulation. Peritoneal T lymphocytes responded to the mitogens less than peripheral blood lymphocytes of the same individual. Supplementation of cell culture with anti-macrophage (anti-CD68) and anti-HLA-DR MoAb brought about a dose-dependent decrease of proliferative peritoneal cell response to ConA. The authors conclude that human peritoneal cell population comprises a high proportion of T lymphocytes and macrophages capable of presenting antigens to peritoneal lymphocytes. High prevalence of memory lymphocytes points to the preparedness of these cells to react with invading antigens most likely of gut bacterial origin.

Rejection of cartilage formed by transplanted allogeneic chondrocytes: evaluation with monoclonal antibodies.

Cellular infiltrates participating in rejection of cartilage formed by transplanted allogeneic rat epiphyseal chondrocytes were evaluated immunohistochemically using a panel of different monoclonal antibodies. One week after transplantation, the grafts were surrounded by numerous class II MHC+ (OX6+, OX17+), CD4+ (W3/25+), and W3/13+ cells as well as some ED1+ monocytes/macrophages. Only a few T (OX19+) and B (HIS14+) cells were present. The number of class II MHC+ cells and ED1+ monocytes/macrophages did not change significantly in the course of rejection whereas the number of CD4+ and W3/13+ cells gradually decreased. On the other hand, there was a significant increase in the number of CD8+ (OX8+) cells. CD8+ cells accumulated close to the transplants and some of them penetrated cartilage matrix suggesting that they might be involved in chondrocyte killing. After 3 months, cartilage was almost completely destroyed and the intensity of infiltrations was markedly decreased. Fibrous connective tissue predominated, however, some class II+ as well as few ED1+, CD4+ and CD8+ cells were still present adjacent to the cartilage remnants. At the time of transplantation, chondrocytes were endowed with RT1.D class II antigen (OX17+), but they did not react with OX6 mAb (monoclonal antibody) recognizing the RT1.B class II molecule. However, after 1 week, some chondrocytes reacted with OX6 mAb and the number of RT1.B positive chondrocytes increased in the course of cartilage rejection.

Skin changes in filarial and non-filarial lymphoedema of the lower extremities.

The pathogenesis of lymphoedema in patients infected with Wuchereria bancrofti or Brugia malayi remains unclear. Lymph stasis and local immunological reactions seem to play the main role. In order to discriminate between the obstructive and immunological effects of the parasite, a comparative histological study of skin specimens obtained from two groups of patients, one with filarial and the other with postsurgical lymphedema of lower extremities, was performed. In both groups patients suffered lymph stasis, in the first due to filariasis, in the other due to removal or irradiation of pelvic lymph nodes. The patients with filarial infection showed hyperproliferation of keratinocytes, focal acantholysis, accumulation of lymphocytes at the epidermo-dermal junction, profuse pericapillary and perivenular mononuclear infiltrations in the dermis marginated granulocytes in capillaries and, in some cases, subepidermal granulocytic infiltrates. There were many dilated initial lymphatics and lymphatic "lakes" between thick collagen fibre bundles. Monoclonal antibody analysis revealed that the most common cells in the infiltrates were macrophages (CD68+). All mononuclear and endothelial cells were HLA-DR+. In contrast, the skin specimens of non-filarial patients revealed only moderate proliferation of keratinocytes, increased numbers of CD1+ epidermal Langerhans cells, moderate pericapillary infiltrates of CD68+, CD4+ and CD8+ cells, and evidently less intensive marking of cells with anti-HLA-DR antibody. There were few initial lymphatics visible. These findings indicate that filarial lymphoedema is complicated by a severe inflammatory component, which is much less expressed in postsurgical lymph stasis.

Normal human immune peritoneal cells: phenotypic characteristics.

Normal human peritoneal cells (PC) collected from patients with calculous cholecystitis without clinically detectable inflammatory changes were characterized morphologically, histochemically and phenotypically by means of monoclonal antibodies. The PC consisted of 45% of monocytes/macrophages (M718 + cells). Thirty-five per cent of PC were esterase-positive and 23% acid phosphatase positive. Forty-five per cent of PC adhered to glass surface. In the lymphocyte population, 2% of CD22 B lymphocytes (M738 +) and 42% CD2 T lymphocytes (M720+) were found. CD4/CD8 ratio was 0.4. There were 8% of Leu7 + cells. The PC did not reveal interleukin 2 (OKT26a +) and transferrin receptors (OKT9 +) on their surface. No blast cells were detected in the PC suspension. Approximately 49% of the PC expressed Ia antigens (OKIa1 +). Two per cent of S100 positive dendritic cells (Z311 +) were found. Peritoneal fluid contained 9% of granulocytes, mostly neutrophils. Two per cent of PC were free mesothelial cells (M717 +). We conclude that human peritoneal cavity contains a cell population significantly differing from that which is present in peripheral blood, which strongly suggests a non-random cell accumulation in the peritoneum. Lack of any activated cells indicates that under normal conditions the peritoneum lavage fluid contains a steady-state population. We conclude that the normal peritoneal fluid cells represent a heterogeneous population capable of reacting to various antigens entering the cavity from the gut.