Long-term effects of low-density lipoprotein apheresis using an automated dextran sulfate cellulose adsorption system. Liposorber Study Group.

The short-term effectiveness of low-density lipoprotein (LDL) apheresis using a dextran sulfate cellulose adsorption column technique was previously examined in a 9-center, 22-week controlled trial in 64 patients with familial hypercholesterolemia (FH) who did not adequately respond to diet and drug therapy. Forty-nine patients (40 treatment, 9 controls) subsequently received LDL apheresis procedures as part of an optional follow-up phase. This study reports on the long-term safety, lipid lowering, and clinical efficacy of LDL apheresis for the 5-year period that includes both the initial controlled study and follow-up phase. During this time, patients received a total of 3,902 treatments of which 3,314 treatments were given during the follow-up phase. Adverse events were infrequent, occurring in 142 procedures (3.6%). Immediate reduction in LDL cholesterol was 76% both in homozygotes and in heterozygotes. Patients with homozygous FH had a progressive decrease in pretreatment LDL cholesterol level along with an increase in high-density lipoprotein (HDL) cholesterol level. There was no appreciable change in pretreatment lipoprotein level over time in heterozygotes. The rate of cardiovascular events during therapy with LDL apheresis and lipid-lowering drugs was 3.5 events per 1,000 patient-months of treatment compared with 6.3 events per 1,000 patient-months for the 5 years before LDL apheresis therapy. These findings support the long-term safety and clinical efficacy of LDL apheresis in patients with heterozygous and homozygous FH who are inadequately controlled with drug therapy.

Increased antibody production in peripheral blood mononuclear cells after plasma exchange therapy in multiple sclerosis.

It has been difficult to study antibody synthesis in vivo following plasma exchange (PE) because of the intra- and extravascular compartmentation of immunoglobulins, and their altered lymphatic flow and catabolism after the procedure. There is fragmentary experimental and clinical evidence that antibody synthesis may increase due to removal of autoregulatory factors by PE. The present study showed increased in vitro spontaneous immunoglobulin (Ig) G and IgM production, as well as increased spontaneous lymphocytic proliferation, after intensive PE in 10 multiple sclerosis (MS) patients. Cytofluorometric control of cultured mononuclear cell populations revealed no change with treatment except for a small increase in B lymphocytes inadequate to account for the increased immunoglobulin production. These findings indicate an activation of the immune system through PE, and suggest the possible need for adjunctive immunosuppression to control antibody production in autoimmune diseases with subacute, relapsing, or chronic courses when PE is employed as a therapeutic modality.

Intrafamily fragment analysis of the T cell receptor beta chain CDR3 region.

The length distributions of the third complementarity determining region (CDR3) of the T cell receptor (TCR) beta chain were determined quantitatively from peripheral blood lymphocytes of healthy donors. RT-PCR products of 26 V beta families and subfamilies were analyzed by an A.L.F. DNA sequencer and Fragment Manager software. We established a normal reference of CDR3 lengths for most of the V beta families and subfamilies. The known range of CDR3 lengths for the V beta chain was expanded to 24 amino acids, and quantitative measurements were made for each fragment length allowing intrafamily CDR3 fragment length comparisons. Importantly, we were able to analyze intrafamily CDR3 fragment distribution without optimizing PCR conditions, thereby circumventing a major obstacle found in intrafamily TCR beta chain comparisons.

Immunologic rebound.

Evidence for accelerated specific antibody (Ab) rebound or overshoot after single or multiple therapeutic plasmapheresis (TP) is fragmentary but suggested by both clinical and experimental evidence. In vitro studies showing increased peripheral blood lymphocyte turnover and total immunoglobulin production after a series of TP without immunosuppression may signify a generalized immunostimulation through removal of regulatory molecules by TP. It is known that IgG class Ab can down regulate B cells by cross linkage of their Ag and Fc receptors. Cyclophosphamide and other cytotoxic immunosuppressive agents effectively delete proliferating lymphocytes. TP could particularly foster deletion of lymphocytes actively mediating autoimmunity, since they would be more readily stimulated to proliferation by removal of Ab or other inhibitory factors than the generally resting normal immune system. This is supported by a relatively greater reduction of autoantibody levels than total immunoglobulin after treatment with TP and cytotoxic immunosuppressives.

Atherosclerotic plaque evolution in the descending thoracic aorta in familial hypercholesterolemic patients. A transesophageal echo study.

We explored the concept that transesophageal echocardiography can be used as a tool to detect, characterize, and study plaque morphology in the descending thoracic aorta. The pattern of atherosclerotic plaques in the descending thoracic aorta in familial hypercholesterolemic (FH) patients was evaluated. Additionally, evolution of plaque characteristics as a result of therapy was analyzed. In a randomized prospective protocol, eight FH patients (five men and three women, aged 23 to 65 years [mean +/- SD, 42 +/- 14 years]) receiving standard therapy (n = 3; baseline low-density lipoprotein [LDL] cholesterol, 222 +/- 71 mg/dL, mean +/- SD) or LDL apheresis (n = 5; baseline LDL cholesterol, 262 +/- 51 mg/dL) were studied. Baseline and follow-up (mean, 12 months) transesophageal echocardiographic studies were performed. Measurements obtained were atherosclerotic plaque area (PA), aortic wall area (WA), total arterial area (TAA), and plaque-to-wall area ratio (PWR). LDL cholesterol decreased in both groups. The greatest severity of plaque was detected at 30 to 35 cm from the incisors (approximately 15 to 20 cm from the aortic arch). The smallest plaques were present at the arch and more distal descending aorta. In the control group, TAA, PA, and PWR did not change significantly (P = NS versus baseline). In the LDL-apheresis group, TAA increased (P < .05 versus baseline), PA decreased in three of five patients (P = NS versus baseline), and PWR fell (P < .05 versus baseline).(ABSTRACT TRUNCATED AT 250 WORDS)

Immune modulation during treatment of systemic sclerosis with plasmapheresis and immunosuppressive drugs.

Immunologic effects of our previously reported treatment protocol were determined in eight systemic sclerosis (SSc) patients treated with plasmapheresis, cyclophosphamide, and prednisone as tolerated. Prior to treatment total serum IgG and circulating lymphocytes including the percentage of B, T, and NK cells were normal. Peripheral blood lymphocytes showed evidence of activation by increased spontaneous proliferation and expression of CD25 on T cells. In addition, CD4+ T cells showed increased expression of both activation (HLA-DR) and maturation (CD29) markers. The percentage of CD8+ T cells was low, resulting in a high CD4:CD8 T cell ratio. During treatment all patients showed clinical improvement. Laboratory testing showed that their NK cells had declined by 81%, B cells declined by 96%, and serum IgG declined by 49%; high titer ANA were abrogated. Total T cells declined by 52%, and the CD4:CD8 ratio fell to normal, due to an almost twofold increase in the percentage of CD8+ T cells. Spontaneous lymphocyte proliferation increased further and was accompanied by increased maturation and activation of both CD4+ and CD8+ T cells, as well as a reduction of immature CD4+ T cells. Among CD8+ T cells the percentage of cytotoxic cells increased as shown by an increase in the CD11b-, CD38+, and CD29+ phenotypes, a finding confirmed on follow-up three-color flow cytometry which showed an increase in activated CD8 cells bearing the cytotoxic CD28 marker. Three-color cytometry also showed that cells with the CD4+CD45RA+CD31+ suppressor inducer phenotype were low and those with the CD4+CD45RO+ CD31- helper inducer phenotype were high in treated SSc patients. Deficient CD4+CD45RA+CD31+ and CD8+ T cell populations suggest an immune regulatory imbalance in SSc which could have led to CD4+ T cell activation and autoimmunity. Depletion of B cells, in conjunction with augmentation of cytotoxic CD8+ T cells through combined therapy, may have diminished the autoimmune response.

Immunomodulation during treatment of polymyositis with plasmapheresis and immunosuppressive drugs.

Immunologic studies were carried out in a patient with polymyositis (PM), who showed increasing muscle strength and decreasing serum creatine phosphokinase levels during 20 weeks of treatment with plasmapheresis in conjunction with prednisone and cyclophosphamide. After an initial rise, serum IgG declined with treatment. Natural killer (NK) lymphocytes were reduced by 74%, B cells by 95%, and T cells by 38%. Spontaneous proliferation of peripheral blood mononuclear cells increased dramatically. Within the CD4+ T cell subset there was increasing maturation as shown by a rise in percent mature (CD29+) cells and reciprocal decline of immature (CD45RA+) cells. At the same time CD4+ T cells became increasingly activated as shown by HLA-DR expression. The percentage of CD8+ T cells increased strongly with treatment, and they showed increased activation and expression of the cytotoxic CD29+ and CD11b- phenotypes. CD8+ T cells exhibiting CD45RA or CD11b+ suppressor phenotypes were overall unchanged; however, on follow-up a proportion of CD8+ cells expressed the activated suppressor effector (CD11b-CD28-) phenotype. In addition to control of PM by the possible deletion of activated autoreactive B and T lymphocyte clones with cyclophosphamide, the activation and maturation of CD4+ T cells during treatment may have downregulated the autoreactive disease process, either through direct antiidiotypic suppression or by induction of the observed increase in cytotoxic and suppressor CD8+ T cells.

Treatment of refractory familial hypercholesterolemia by low-density lipoprotein apheresis using an automated dextran sulfate cellulose adsorption system. The Liposorber Study Group.

A subgroup of patients with familial hypercholesterolemia (FH) respond inadequately to standard diet and drug therapy, and are therefore at high risk for the premature development or progression of coronary artery disease. This study evaluated low-density lipoprotein (LDL) cholesterol and lipoprotein (a) removal in a multicenter, controlled trial with a new LDL apheresis procedure (Liposorber LA-15 System). The study comprised patients with FH who had not responded adequately to diet and maximal drug therapy. There were 54 patients with heterozygous FH (45 randomized to treatment and 9 control subjects) and 10 with homozygous FH (all of whom received LDL apheresis). The study included three 6-week treatment phases and a 4-week rebound phase. Treatments were administered at 7- to 14-day intervals. Mean acute reductions in LDL cholesterol were 76% in heterozygous FH patients and 81% in homozygous ones. Time-averaged levels of LDL cholesterol were reduced 41% (243 to 143 mg/dl) in heterozygous FH patients and 53% (447 to 210 mg/dl) in homozygous ones. The substantial acute reduction of lipoprotein (a) (means: 65%, heterozygous FH; 68%, homozygous FH) has not been reported with other therapies. The Liposorber LA-15 System represents an important therapeutic option in FH patients who respond inadequately to diet and drug therapy.

Chronic arsenic poisoning masquerading as Landry-Guillain-Barré syndrome.

Acute arsenic intoxication may present as Landry-Guillain-Barré syndrome because of similarities in clinical symptoms involving the gastrointestinal tract, weakness, and sensory symptoms. Electrodiagnostic findings may be similar with demyelinating changes predominating early in both diseases. A case is presented of repeated arsenic poisoning over two years misdiagnosed as Landry-Guillain-Barré syndrome. Proximal F-loop latency (M-wave latency at wrist + F-wave latency at wrist - 2 M-wave latency at axilla) helped to establish the correct diagnosis. Serial electrodiagnostic studies were done documenting the evolution of chronic repeated arsenic poisoning from a picture showing demyelination to one with severe axonal loss.

Immunologic effects of plasmapheresis synchronized with pulse cyclophosphamide in systemic lupus erythematosus.

Extensive immunologic evaluation was made of a patient with severe systemic lupus erythematosus (SLE) undergoing 6 cycles of plasmapheresis combined with pulsed cyclophosphamide therapy. Clinical remission ensued accompanied by normalization of levels of circulating autoantibodies, immune complexes, complement proteins, interleukin 2 (IL-2) and IL-2 receptor. High spontaneous peripheral blood lymphocyte proliferation fluctuated widely with plasmapheresis, but was consistently reduced after cyclophosphamide. Circulating B cells fell by 85% and remained low at one year, despite recovery of the serum IgG level. Circulating T cells declined by 48%, chiefly in the immunologically naive CD4+CD45R+ T cell subset. This was associated with the emergence of a CD8+DR+CDw29+ T cell subset signifying immunologically mature, activated cytotoxic/suppressive T cells, which might have served to control the autoreactive B and T cell populations. Pulsed cyclophosphamide synchronized with plasmapheresis profoundly affected the immune system of our patient. The association of these immunological changes with clinical recovery warrants further investigation of this new therapeutic approach in SLE.

Plasmapheresis and subsequent pulse cyclophosphamide versus pulse cyclophosphamide alone in severe lupus: design of the LPSG trial. Lupus Plasmapheresis Study Group (LPSG).

A group of clinics are collaborating in the Lupus Plasmapheresis Study Group (LPSG) to investigate whether repeated plasmapheresis prior to pulse cyclophosphamide improves the therapeutical results in severe systemic lupus erythematosus (SLE). The underlying rationale is the hypothesis that plasmapheresis 1) eliminates pathogenic autoantibodies and immune complexes and 2) induces compensatory lymphocyte activation via feedback mechanisms between circulating antibodies and their respective clones ("antibody rebound"). It should be possible to utilize this enhanced activity for increased clonal deletion if pulse cyclophosphamide is applied shortly after plasmapheresis. Accordingly, in a randomized study, the LPSG will be comparing the repeated application of pulse cyclophosphamide alone with a treatment involving repeated plasmapheresis prior to the cyclophosphamide pulses in severe SLE. A third arm of the study will be gathering experience with a more intensified procedure. This overview summarizes the most important details of the planned study.

Increased proliferation of blood mononuclear cells after plasmapheresis treatment of patients with demyelinating disease.

Induction of lymphocytic proliferation has been postulated to be a mechanism whereby plasmapheresis may enhance the action of cytotoxic immunosuppressive drugs. This study found increased spontaneous proliferation of peripheral blood mononuclear cells following intensive plasmapheresis treatment of patients with multiple sclerosis (MS) or Guillain-Barré syndrome (GBS). The increased proliferative response was reduced below baseline in four of six MS patients who received subsequent immunoglobulin intravenous (IGIV) and pulsed cyclophosphamide therapy, but not in three MS patients receiving IGIV alone. In five GBS patients with low baseline proliferation, proliferation also increased after plasmapheresis. High baseline proliferation found in three GBS patients may have reflected antecedent infection, since it fell during plasmapheresis in the two patients in whom it was measured. Plasmapheresis could possibly augment the effectiveness of cytotoxic drugs in controlling autoimmunity by inducing lymphocytes to proliferate, thereby making them more susceptible to drug action.

Double-blind study of true vs. sham plasma exchange in patients treated with immunosuppression for acute attacks of multiple sclerosis.

We enrolled 116 patients in a multicenter, randomized, double-blind controlled trial of an 8-week course of 11 plasma exchange (PE) treatments in exacerbations of MS. The control group received sham PE, and both groups received identical treatment with IM ACTH and oral cyclophosphamide. Serum IgG decreased in the PE and sham treatment groups by 76% versus 22% by treatment 5, and by 64% versus 14% by treatment 11. PE also produced significant reductions in IgA, IgM, C3, and fibrinogen. PE patients had moderately enhanced improvement at 2 weeks relative to the sham group. PE patients with relapsing/remitting disease had significantly enhanced improvement at 4 weeks and there was also an increased improvement at 12 months, although this latter effect disappeared when we analyzed relapsing/remitting patients as a separate subgroup. Life table analysis showed the median time to recover preattack disability status was shorter in PE- than in sham-treated relapsing/remitting patients (4 vs. 13 weeks), a result confirmed by raw disability status scores in which there was recovery to their average preattack disability score by 3 months. PE given with ACTH plus cyclophosphamide enhances recovery from an exacerbation of disease in relapsing/remitting patients, although we observed no clear long-term benefits.

Remission of hyper-IgE syndrome treated with plasmapheresis and cytotoxic immunosuppression.

A patient with a hyper-IgE syndrome was treated with 60 plasmaphereses over a period of 2 years in conjunction with cytotoxic immunosuppressive drug therapy. During this time her severe dermatitis of 8 years' duration became almost completely inactive, and her circulating IgE level was reduced by 73%. An elevated pretreatment ratio of CD4+/CD8+ T lymphocytes fell to subnormal. The beneficial results of treatment may be attributed to the reduction of lymphocyte populations responsible for IgE production by the combined action of plasmapheresis and of cytotoxic drugs as well as the direct effect of removal of circulating IgE and possibly IgE-potentiating factors.

Reduction of low-density lipoproteins with dextran sulfate in patients with familial hypercholesterolemia.

A novel on-line system for the selective precipitation of low-density lipoprotein (LDL) using dextran sulfate has been devised and tested in four patients with heterozygous familial hypercholesterolemia (type II). The mean pretreatment serum cholesterol was 410 mg/dl. Plasma was generated by membrane filtration and LDL and VLDL (very-low-density lipoprotein) were completely precipitated with 10-35 mg% dextran sulfate (Mr 5,000) in the presence of 55 mM Ca2+. The precipitate was removed by filtration and the excess Ca2+ by dialysis. For 41 procedures the mean reduction of plasma solutes was LDL + VLDL 65%, HDL 23%, fibrinogen 19%, albumin 15%, IgG 20%, IgA 19%, IgM 24%. We conclude that dextran sulfate precipitation is an effective method for selective on-line removal of LDL from plasma.

Treatment of hypercholesterolemia by precipitation of lipoproteins with dextran sulfate.

An on-line continuous system for the selective precipitation of low-density lipoproteins (LDL) and very low-density lipoproteins (VLDL) has been devised and tested. This system conserves high-density lipoproteins (HDL) and other plasma macromolecules. LDL and VLDL are precipitated from plasma using 10-35 mg/dl dextran sulfate (Mr 5,000) in the presence of 55 mM calcium with a reduced concentration of monovalent cations. The plasma is obtained by membrane filtration of whole blood using the COBE Centry TPE System (Cobe Laboratories Inc, Lakewood, Co.). The precipitated LDL plus VLDL is removed by filtration, and the electrolytes are restored by dialysis. The plasma minus LDL plus VLDL is then returned to the patient. Four patients with heterozygous familial hypercholesterolemia (type II) were treated 70 times. The mean pretreatment serum cholesterol was 383 mg/dl. The mean reductions in plasma components were: LDL plus VLDL 63%; HDL 27%; fibrinogen 19%; albumin 15%; IgG 20%; IgA 19%; IgM 25%; C3 30%; and C4 27%. The cholesterol returned to near normal values in approximately 2 weeks after each treatment. Four normal volunteers were each treated one time. These individuals had a mean pretreatment serum cholesterol of 201 mg/dl. The mean reduction in plasma components were: LDL plus VLDL 70%; HDL 27%; fibrinogen 24%; albumin 14%; IgG 18%; IgA 17%; IgM 20%; C3 27%; C4 22%; C3 proactivator 12%; alpha 1-antitrypsin 17%; ceruloplasma 17%; transferrin 18%; alpha 2-macroglobulin 17%; and orosomucoid 13%. It is our conclusion that dextran sulfate precipitation is an effective on-line means of selectively removing LDL plus VLDL from plasma while conserving HDL and other plasma macromolecules.(ABSTRACT TRUNCATED AT 250 WORDS)

Induction of in vitro autoimmune responses by mononuclear blood cells in Hashimoto's thyroiditis.

T cell proliferative responses were generated in vitro in lymphocytes from Hashimoto's thyroiditis (HT) patients using antigen presenting cells (APC) separated by fibronectin adherence and culture in serum free medium (SFM). HT cells were found to respond to the heterologous antigen, keyhole limpet haemocyanin (KLH), as well as to the homologous antigens thyroglobulin (Tg), myelin basic protein (MBP), myoglobin (Mb), and sarcolemma (Sl). Positive responses were dependent upon gamma-interferon, interleukin 1, and interleukin 2. The ratio of HT T helper/inducer cells to T suppressor/cytotoxic cells was elevated at the inception of culture, and further increased during the 9 day culture period. Our data show a generalized defect in self-tolerance by HT T cells which may have been revealed by preincubation of antigen with restricted numbers of APC and subsequent culture in SFM.