Consistency of a dialyzable leucocyte extract manufactured at GMP facilities by nuclear magnetic resonance spectroscopy.

The present work describes the development and validation of a first report including several non-invasive NMR schemes to identify parameters as local chemical environments, homo- and heteronuclear site-specific spin correlations, diffusion coefficient-dependent polydispersity indexes and quantification of identified peptide entities that composes a commercial human Dialyzable Leucocyte Extract (DLE), Transferon, an oral liquid formulation of low-molecular-weight peptides. The above parameters were useful indicators to verify reproducibility, consistency and homogeneity among the DLE batches manufactured at Good Manufacturing Practice (GMP) facilities and for batch-releasing purposes in a quality control laboratory. The results showed that peptide identity of the DLE is represented with both high reproducible one-dimensional proton spectra and diffusion coefficient distributions that predicts in turn a weight-average molecular weight of around 6.7-7.4 kDa and a mean polydispersity index of 1.13. The obtained NMR peptide fingerprint of the analyzed DLE allowed to i) confirm its structural homogeneity by line-shape analysis, ii) identify and quantify its peptide content within the total solution with qNMR methods iii) to confirm the robustness of the technique as a feasible alternative for routine analysis of Natural or non-Natural Complex Drugs, such as DLEs.

Anti-inflammatory effect of omega unsaturated fatty acids and dialysable leucocyte extracts on collagen-induced arthritis in DBA/1 mice.

Rheumatoid arthritis is a disabling autoimmune disease with a high global prevalence. Treatment with disease-modifying anti-arthritic drugs (DIMARDs) has been routinely used with beneficial effects but with adverse long-term consequences; novel targeted biologics and small-molecule inhibitors are promising options. In this study, we investigated whether purified omega unsaturated fatty acids (ω-UFAs) and dialysable leukocyte extracts (DLEs) prevented the development of arthritis in a model of collagen-induced arthritis (CIA) in mice. We also investigated whether the transcription factor NF-κB and the NLRP3 inflammasome were involved in the process and whether their activity was modulated by treatment. The development of arthritis was evaluated for 84 days following treatment with nothing, dexamethasone, DLEs, docosahexaenoic acid, arachidonic acid, and oleic acid. Progression of CIA was monitored by evaluating clinical manifestations, inflammatory changes, and histological alterations in the pads' articular tissues. Both DLEs and ω-UFAs led to an almost complete inhibition of the inflammatory histopathology of CIA and this was concomitant with the inhibition of NF-kB and the inhibition of the activation of NLRP3. These data suggest that ω-UFAs and DLEs might have NF-κB as a common target and that they might be used as ancillary medicines in the treatment of arthritis.

Valproic acid promotes a decrease in mycobacterial survival by enhancing nitric oxide production in macrophages stimulated with IFN-γ.

Tuberculosis is one of the leading causes of mortality worldwide, it is caused by Mycobacterium tuberculosis (Mtb), a bacteria that employs several strategies to evade the host immune response. For instance, Mtb interferes with the overexpression of class II transactivator (CIITA) in macrophages exposed to IFN-γ by inhibiting histone acetylation at its promoter, which can be reverted by the histone deacetylase inhibitor (HDACi) sodium butyrate. In this work, we evaluated whether a different HDACi, valproic acid (VPA), could revert the inhibition of gene expression induced by Mtb. J774 macrophages treated with VPA and IFN-γ unexpectedly induced a higher expression of the inducible nitric oxide synthase and a higher production of nitric oxide when exposed to the 19 kDa lipoprotein of Mtb or the whole bacteria. However, VPA was unable to revert the inhibition of CIITA expression induced by the 19 kDa lipoprotein of Mtb. Finally, macrophages infected with Mtb and treated with VPA and IFN-γ showed a significant reduction in intracellular bacteria. Our findings suggest a new therapeutic potential of VPA for the treatment of tuberculosis.

Dialyzable leukocyte extracts activate TLR-2 on monocytes.

Dialyzable leukocyte extracts (DLE) transfer specific cell-mediated immune responses from sensitized donors to non-immune recipients. In addition, DLE have several immunomodulatory effects and are used for the treatment of several infectious and non-infectious diseases. Previous studies showed that human DLE obtained from virus-infected leukocytes and bovine DLE decrease the production of the pro-inflammatory cytokine TNF-alpha in response to bacterial lipopolysaccharide, in vitro and in vivo. In the present work, we inquire as to whether DLE from uninfected human leukocytes have the ability to regulate cytokine production in peripheral blood mononuclear cells (PBMC) in vitro. We observed that PBMC from healthy individuals were able to produce TNF-alpha, IL-12 and IL-10 after stimulation with DLE. Moreover, we identified monocytes as the main cell population that produced TNF-alpha after DLE stimulation. Interestingly, we found that DLE contain unidentified ligands that activate Toll-like receptor (TLR)-2. Finally, we observed that DLE directly activated monocytes through TLR-2. These results reveal a new biological activity of DLE, and suggest that part of the immunomodulatory properties of DLE could be attributed to TLR-2 activation on monocytes and to the induction of a pro-inflammatory environment that is crucial for control of infectious diseases.

Effect of selective serotonin reuptake inhibitors and immunomodulator on cytokines levels: an alternative therapy for patients with major depressive disorder.

Major depressive disorder (MDD) is a psychiatric illness that presents as a deficit of serotonergic neurotransmission in the central nervous system. MDD patients also experience alterations in cortisol and cytokines levels. Treatment with selective serotonin reuptake inhibitors (SSRIs) is the first-line antidepressant regimen for MDD. The aim of this study was to determine the effect of a combination of SSRIs and an immunomodulator-human dialyzable leukocyte extract (hDLE)-on cortisol and cytokines levels. Patients received SSRIs or SSRIs plus hDLE. The proinflammatory cytokines IL-1 ß , IL-2, and IFN- γ ; anti-inflammatory cytokines IL-13 and IL-10; and 24-h urine cortisol were measured at weeks (W) 0, 5, 20, 36, and 52 of treatment. The reduction in cortisol levels in the SSRI-treated group was 30% until W52, in contrast, the combined treatment induced a 54% decrease at W36. The decline in cortisol in patients who were treated with SSRI plus hDLE correlated with reduction of anti-inflammatory cytokines and increases levels of proinflammatory cytokines at the study conclusion. These results suggest that the immune-stimulating activity of hDLE, in combination with SSRIs, restored the pro- and anti-inflammatory cytokine balance and cortisol levels in depressed patients versus those who were given SSRIs alone.

Effect of glycine on the immune response of the experimentally diabetic rats.

BACKGROUND: Hyperglycemia induces protein glycation, disturbing its function, additionally, the glycated products (AGEs) induce by themselves proinflammatory cytokine release that are responsible for insulin resistance. Glycine has been successfully used in diabetic patients to competitively reduce hemoglobin glycation. OBJECTIVES: To assess hyperglycemia impact on the immune response and to evaluate if it is possible to reverse it by means of glycine administration. MATERIAL AND METHODS: Streptozotocin-induced diabetic rats, with and without glycine administration were challenged with sheep red blood cells, and specific antibody producing cells were accounted. Normal rats were challenged as controls. RESULTS: Induced diabetes modifies significantly the humoral immune response capacity versus sheep red blood cells. Also, glycine administration prevents against this deleterious effect. CONCLUSIONS: Glycine could be an important therapeutic resource among diabetics to avoid the characteristic immunodeficiencies of this disease.

[Psychological, immunological, and endocrinological implications of atopic dermatitis]. / Implicaciones psicológicas, inmunológicas y endocrinológicas en la dermatitis atópica.

BACKGROUND: The atopic dermatitis is a chronic disease with immunological alterations, neuro-immune-endocrine implications and repercussions in the psyche. OBJECTIVE: To determine the effectiveness of the use of psychological support in the treatment of atopic dermatitis. MATERIAL AND METHODS: This study included 20 patients (from 15 to 50 years old) with moderated and severe atopic dermatitis based on the Hanifin and Rafka modified criteria. They were classified in three groups of treatment: A) 10 patients with transfer factor and psychological support, B) 10 with transference factor without psychological support, C) 10 healthy individuals as a control group. At the beginning and ending of the analysis the degree of severity of the illness was determined using the SCORAD scale. RESULTS: There was a more statistically significant clinical improvement on group A than in group B, besides a decrease in anxiety and depression, at a laboratory level including hormonal behavior, without meaningful changes. CONCLUSIONS: It was demonstrated that the treatment with immune-modulator and psychological support on the moderated and severe atopic dermatitis improves the clinical and immunological evolution of this illness.

Seguridad y eficacia en el tratamiento de la dermatitis atópica severa con ciclosporina A y factor de transferencia / The study of two therapeutic alternatives, Transfer Factor (TF) and Cyclosporin A in atopic dermatitis

Antecedentes. La dermatitis atópica es una enfermedad crónica de la piel que se presenta en pacientes con historia personal o familiar de asma y rinitis alérgica vinculada con la activación de un grupo específico de genes. Existen casos en los que la respuesta al tratamiento convencional no es la esperada, a los que se considera como refractarios. Para este tipo de pacientes se elaboró un estudio en donde se comparan dos alternativas terapéuticas: factor de transferencia (FT) y la ciclosporina A (C y A). Material y método. Se estudiaron pacientes con diagnóstico de dermatitis atópica severa refractaria a cualquier tratamiento, que ingresaron al servicio de alergia del Hospital Regional Lic. Adolfo López Mateos, ISSSTE, entre los meses de julio de 1997 a septiembre de 1998. Se distribuyeron por sorteo en dos grupos: el primero a quienes se indicó ciclosporina A a la dosis de 4 mg/kg/día durante tres meses, con vigilancia de la función renal y hepática en forma mensual, y en las cifras de tensión arterial dos veces por semana. El grupo dos fue tratado con factor de transferencia a la dosis de una unidad cada tercer día durante la primera semana, dos unidades por semana durante tres semanas y, finalmente, una unidad mensual hasta completar seis meses de tratamiento. En ambos grupos se realizó valoración clínica e inmunológica (eosinófilos, IgE, subpoblaciones linfocitarias) inicial y final. Resultados. Se incluyeron en el estudio ocho pacientes del grupo A y 12 en el grupo B. Ambos presentaron mejoría que se cuantificó como estadísticamente significativa en la reducción de eosinófilos periféricos, sin diferencia estadística entre ellos. Ninguno tuvo modificaciones en las cifras de IgE total. La ciclosporina A presentó disminución de las concentraciones de CD8+, ambos con una p<0.05. Los dos grupos tuvieron mejoría clínica cuantificada como estadísticamente significativa; sin embargo, no hubo diferencias entre ambos grupos con una p>0.05. La tolerancia a los tratamientos fue adecuada, en ningún caso fue necesario suspender el tratamiento. Sólo sobrevino en un caso que recibió ciclosporina A, hipertricosis en tres pacientes y cefalea en otro. Conclusión. Ambas opciones mostraron beneficio en el tratamiento de pacientes con dermatitis atópica severa refractaria, con mejoría clínica e inmunológica similar...