Use of Cells, Supplements, and Peptides as Therapeutic Strategies for Modulating Inflammation after Spinal Cord Injury: An Update.

Spinal cord injury is a traumatic lesion that causes a catastrophic condition in patients, resulting in neuronal deficit and loss of motor and sensory function. That loss is caused by secondary injury events following mechanical damage, which results in cell death. One of the most important events is inflammation, which activates molecules like proinflammatory cytokines (IL-1ß, IFN-γ, and TNF-α) that provoke a toxic environment, inhibiting axonal growth and exacerbating CNS damage. As there is no effective treatment, one of the developed therapies is neuroprotection of the tissue to preserve healthy tissue. Among the strategies that have been developed are the use of cell therapy, the use of peptides, and molecules or supplements that have been shown to favor an anti-inflammatory environment that helps to preserve tissue and cells at the site of injury, thus favoring axonal growth and improved locomotor function. In this review, we will explain some of these strategies used in different animal models of spinal cord injury, their activity as modulators of the immune system, and the benefits they have shown.

Gaucher Disease: Identification and Novel Variants in Mexican and Spanish Patients.

BACKGROUND: Gaucher disease (GD) is the most prevalent lysosomal storage disorder, affecting all ethnic groups, although its prevalence is higher in Ashkenazi Jewish populations. Three clinical forms of GD have been described: Type 1 non-neuronopathic, type 2 acute neuronopathic, and type 3 subacute neuronopathic. An autosomal recessive disorder is caused by variants in the human glucocerebrosidase gene (GBA; MIM*606463) located on chromosome 1q21, resulting from deficit or lack of activity of the ß-glucocerebrosidase enzyme, leading to the accumulation of glucocerebroside substrate in the cells of the macrophage-monocyte system. The aim was to determine variants in Mexican and Spanish populations with GD. METHODS: We report the molecular analysis by a direct automatic method sequenced of both chains of the GBA gene, in 69 Mexican and 369 Spanish patients with GD. RESULTS: We detected 75 variants with pathogenic or likely pathogenic effect and, identified 3 new variants c.408_412del/p.Asn136Lysfs*15; c.820G>A/p.Glu274Lys and c.1058T>G/p*. The most frequent variants were c.1448T>C/p.Leu483Pro/L444P and c.1226A>G/p.Asn409Ser/N370S. The detected genotypes were compared with data from both GD registries to define similarities and differences in both populations. CONCLUSIONS: We defined the variant profile in patients with GD in a Mexican and a Spanish population and compared them. The screening permitted the detection of common variants and the report of three new variants, in addition to a variant associated with Parkinson disease but not with GD. Since molecular diagnosis has considerable predictive value in GD, it is important to study the genotype-phenotype correlations, establishing the severity of the variant.

Quinacrine, an Antimalarial Drug with Strong Activity Inhibiting SARS-CoV-2 Viral Replication In Vitro.

Quinacrine (Qx), a molecule used as an antimalarial, has shown anticancer, antiprion, and antiviral activity. The most relevant antiviral activities of Qx are related to its ability to raise pH in acidic organelles, diminishing viral enzymatic activity for viral cell entry, and its ability to bind to viral DNA and RNA. Moreover, Qx has been used as an immunomodulator in cutaneous lupus erythematosus and various rheumatological diseases, by inhibiting phospholipase A2 modulating the Th1/Th2 response. The aim of this study was to evaluate the potential antiviral effect of Qx against denominated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Vero E6 cells. The cytotoxicity of Qx in Vero E6 cells was determined by the MTT assay. Afterwards, Vero E6 cells were infected with SARS-CoV-2 at different multiplicities of infections (MOIs) of 0.1 and 0.01 in the presence of Qx (0-30 µM) to determinate the half maximal effective concentration (EC50). After 48 h, the effect of Qx against SARS-CoV-2 was assessed by viral cytotoxicity and viral copy numbers, the last were determined by digital real-time RT-PCR (ddRT-PCR). Additionally, electron and confocal microscopy of Vero E6 cells infected and treated with Qx was studied. Our data show that Qx reduces SARS-CoV-2 virus replication and virus cytotoxicity, apparently by inhibition of viral ensemble, as observed by ultrastructural images, suggesting that Qx could be a potential drug for further clinical studies against coronavirus disease 2019 (COVID-19) infection.

Neuroprotective effect of immunomodulatory peptides in rats with traumatic spinal cord injury.

Several therapies have shown obvious effects on structural conservation contributing to motor functional recovery after spinal cord injury (SCI). Nevertheless, neither strategy has achieved a convincing effect. We purposed a combined therapy of immunomodulatory peptides that individually have shown significant effects on motor functional recovery in rats with SCI. The objective of this study was to investigate the effects of the combined therapy of monocyte locomotion inhibitor factor (MLIF), A91 peptide, and glutathione monoethyl ester (GSH-MEE) on chronic-stage spinal cord injury. Female Sprague-Dawley rats underwent a laminectomy of the T9 vertebra and a moderate contusion. Six groups were included: sham, PBS, MLIF + A91, MLIF + GSH-MEE, A91 + GSH-MEE, and MLIF + A91 + GSH-MEE. Two months after injury, motor functional recovery was evaluated using the open field test. Parenchyma and white matter preservation was evaluated using hematoxylin & eosin staining and Luxol Fast Blue staining, respectively. The number of motoneurons in the ventral horn and the number of axonal fibers were determined using hematoxylin & eosin staining and immunohistochemistry, respectively. Collagen deposition was evaluated using Masson's trichrome staining. The combined therapy of MLIF, A91, and GSH-MEE greatly contributed to motor functional recovery and preservation of the medullary parenchyma, white matter, motoneurons, and axonal fibres, and reduced the deposition of collagen in the lesioned area. The combined therapy of MLIF, A91, and GSH-MEE preserved spinal cord tissue integrity and promoted motor functional recovery of rats after SCI. This study was approved by the National Commission for Scientific Research on Bioethics and Biosafety of the Instituto Mexicano del Seguro Social under registration number R-2015-785-116 (approval date November 30, 2015) and R-2017-3603-33 (approval date June 5, 2017).

Motor Recovery after Chronic Spinal Cord Transection in Rats: A Proof-of-Concept Study Evaluating a Combined Strategy.

BACKGROUND: The chronic phase of Spinal Cord (SC) injury is characterized by the presence of a hostile microenvironment that causes low activity and a progressive decline in neurological function; this phase is non-compatible with regeneration. Several treatment strategies have been investigated in chronic SC injury with no satisfactory results. OBJECTIVE- In this proof-of-concept study, we designed a combination therapy (Comb Tx) consisting of surgical glial scar removal plus scar inhibition, accompanied with implantation of mesenchymal stem cells (MSC), and immunization with neural-derived peptides (INDP). METHODS: This study was divided into three subsets, all in which Sprague Dawley rats were subjected to a complete SC transection. Sixty days after injury, animals were randomly allocated into two groups for therapeutic intervention: control group and animals receiving the Comb-Tx. Sixty-three days after treatment we carried out experiments analyzing motor recovery, presence of somatosensory evoked potentials, neural regeneration-related genes, and histological evaluation of serotoninergic fibers. RESULTS: Comb-Tx induced a significant locomotor and electrophysiological recovery. An increase in the expression of regeneration-associated genes and the percentage of 5-HT+ fibers was noted at the caudal stump of the SC of animals receiving the Comb-Tx. There was a significant correlation of locomotor recovery with positive electrophysiological activity, expression of GAP43, and percentage of 5-HT+ fibers. CONCLUSION: Comb-Tx promotes motor and electrophysiological recovery in the chronic phase of SC injury subsequent to a complete transection. Likewise, it is capable of inducing the permissive microenvironment to promote axonal regeneration.

Release of interleukin-10 and neurotrophic factors in the choroid plexus: possible inductors of neurogenesis following copolymer-1 immunization after cerebral ischemia.

Copolymer-1 (Cop-1) is a peptide with immunomodulatory properties, approved by the Food and Drug Administration of United States in the treatment of multiple sclerosis. Cop-1 has been shown to exert neuroprotective effects and induce neurogenesis in cerebral ischemia models. Nevertheless, the mechanism involved in the neurogenic action of this compound remains unknown. The choroid plexus (CP) is a network of cells that constitute the interphase between the immune and central nervous systems, with the ability to mediate neurogenesis through the release of cytokines and growth factors. Therefore, the CP could play a role in Cop-1-induced neurogenesis. In order to determine the participation of the CP in the induction of neurogenesis after Cop-1 immunization, we evaluated the gene expression of various growth factors (brain-derived neurotrophic factor, insulin-like growth factor 1, neurotrophin-3) and cytokines (tumor necrosis factor alpha, interferon-gamma, interleukin-4 (IL-4), IL-10 and IL-17), in the CP at 14 days after ischemia. Furthermore, we analyzed the correlation between the expression of these genes and neurogenesis. Our results showed that Cop-1 was capable of stimulating an upregulation in the expression of the genes encoding for brain-derived neurotrophic factor, insulin-like growth factor 1, neurotrophin-3 and IL-10 in the CP, which correlated with an increase in neurogenesis in the subventricular and subgranular zone. As well, we observed a downregulation of IL-17 gene expression. This study demonstrates the effect of Cop-1 on the expression of growth factors and IL-10 in the CP, in the same way, presents a possible mechanism involved in the neurogenic effect of Cop-1.

The Severity of Spinal Cord Injury Determines the Inflammatory Gene Expression Pattern after Immunization with Neural-Derived Peptides.

Previous studies revealed that the intensity of spinal cord injury (SCI) plays a key role in the therapeutic effects induced by immunizing with neural-derived peptides (INDP), as severe injuries abolish the beneficial effects induced by INDP. In the present study, we analyzed the expression of some inflammation-related genes (IL6, IL12, IL-1ß, IFNÉ£, TNFα, IL-10, IL-4, and IGF-1) by quantitative PCR in rats subjected to SCI and INDP. We investigated the expression of these genes after a moderate or severe contusion. In addition, we evaluated the effect of INDP by utilizing two different peptides: A91 and Cop-1. After moderate injury, both A91 and Cop-1 elicited a pattern of genes characterized by a significant reduction of IL6, IL1ß, and TNFα but an increase in IL10, IL4, and IGF-1 expression. There was no effect on IL-12 and INFÉ£. In contrast, the opposite pattern was observed when rats were subjected to a severe spinal cord contusion. Immunization with either peptide caused a significant increase in the expression of IL-12, IL-1ß, IFNÉ£ (pro-inflammatory genes), and IGF-1. There was no effect on IL-4 and IL-10 compared to controls. After a moderate SCI, INDP reduced pro-inflammatory gene expression and generated a microenvironment prone to neuroprotection. Nevertheless, severe injury elicits the expression of pro-inflammatory genes that could be aggravated by INDP. These findings correlate with our previous results demonstrating that severe injury inhibits the beneficial effects of protective autoimmunity.

Activation of human gonadotropin-releasing hormone receptor promotes down regulation of ARHGAP18 and regulates the cell invasion of MDA-MB-231 cells.

The Gonadotropin-Releasing Hormone Receptor (GnRHR) is expressed mainly in the gonadotrope membrane of the adenohypophysis and its natural ligand, the Gonadotropin-Releasing Hormone (GnRH), is produced in anterior hypothalamus. Furthermore, both molecules are also present in the membrane of cells derived from other reproductive tissues such as the breast, endometrium, ovary, and prostate, as well as in tumors derived from these tissues. The functions of GnRH receptor and its hormone in malignant cells have been related with the decrease of proliferation and the invasiveness of those tumors however, little is known about the molecules associated with the signaling pathways regulated by both molecules in malignant cells. To further analyze the potential mechanisms employed by the GnRHR/GnRH system to reduce the tumorigenesis of the highly invasive breast cancer cell line MDA-MB-231, we performed microarrays experiments to evaluated changes in genes expression and validate these modifications by functional assays. We show that activation of human GnRHR is able to diminish the expression and therefore functions of the Rho GTPase-Activating Protein 18 (ARHGAP18). Decrease of this GAP following GnRHR activation, correlates to the higher of cell adhesion and also with reduction of tumor cell invasion, supporting the notion that GnRHR triggers intracellular signaling pathways that acts through ARHGAP18. On the contrary, although a decline of cellular proliferation was observed during GnRHR activation in MDA-MB-231, this was independent of ARHGAP18 showing the complex system in which is involved the signaling pathways regulated by the GnRHR/GnRH system.

Immunization with neural derived peptides plus scar removal induces a permissive microenvironment, and improves locomotor recovery after chronic spinal cord injury.

BACKGROUND: Immunization with neural derived peptides (INDP) as well as scar removal-separately-have shown to induce morphological and functional improvement after spinal cord injury (SCI). In the present study, we compared the effect of INDP alone versus INDP with scar removal on motor recovery, regeneration-associated and cytokine gene expression, and axonal regeneration after chronic SCI. Scar removal was conducted through a single incision with a double-bladed scalpel along the stump, and scar renewal was halted by adding α,α'-dipyridyl. RESULTS: During the chronic injury stage, two experiments were undertaken. The first experiment was aimed at testing the therapeutic effect of INDP combined with scar removal. Sixty days after therapeutic intervention, the expression of genes encoding for TNFα, IFNγ, IL4, TGFß, BDNF, IGF1, and GAP43 was evaluated at the site of injury. Tyrosine hydroxylase and 5-hydroxytryptamine positive fibers were also studied. Locomotor evaluations showed a significant recovery in the group treated with scar removal + INDP. Moreover; this group presented a significant increase in IL4, TGFß, BDNF, IGF1, and GAP43 expression, but a decrease of TNFα and IFNγ. Also, the spinal cord of animals receiving both treatments presented a significant increase of serotonergic and catecholaminergic fibers as compared to other the groups. The second experiment compared the results of the combined approach versus INDP alone. Rats receiving INDP likewise showed improved motor recovery, although on a lesser scale than those who received the combined treatment. An increase in inflammation and regeneration-associated gene expression, as well as in the percentage of serotonergic and catecholaminergic fibers was observed in INDP-treated rats to a lesser degree than those in the combined therapy group. CONCLUSIONS: These findings suggest that INDP, both alone and in combination with scar removal, could modify the non-permissive microenvironment prevailing at the chronic phase of SCI, providing the opportunity of improving motor recovery.

Cytokine and Growth Factor Activation In Vivo and In Vitro after Spinal Cord Injury.

Spinal cord injury results in a life-disrupting series of deleterious interconnected mechanisms encompassed by the primary and secondary injury. These events are mediated by the upregulation of genes with roles in inflammation, transcription, and signaling proteins. In particular, cytokines and growth factors are signaling proteins that have important roles in the pathophysiology of SCI. The balance between the proinflammatory and anti-inflammatory effects of these molecules plays a critical role in the progression and outcome of the lesion. The excessive inflammatory Th1 and Th17 phenotypes observed after SCI tilt the scale towards a proinflammatory environment, which exacerbates the deleterious mechanisms present after the injury. These mechanisms include the disruption of the spinal cord blood barrier, edema and ion imbalance, in particular intracellular calcium and sodium concentrations, glutamate excitotoxicity, free radicals, and the inflammatory response contributing to the neurodegenerative process which is characterized by demyelination and apoptosis of neuronal tissue.

Anti-inflammatory, antioxidant and anti-acetylcholinesterase activities of Bouvardia ternifolia: potential implications in Alzheimer's disease.

Bouvardia ternifolia has been used medicinally to treat inflammation. In the present study, we investigate the anti-Alzheimer's potential effect of the hydroalcoholic extract of B. ternifolia through evaluation of anti-inflammatory and antioxidant activities, quantification of the percentage inhibition of acetylcholinesterase activity, protection effect against ß-amyloid fibrillar-induce neurotoxicity, and the identification of the main constituents. Our results show that B. ternifolia extract and ethyl acetate fraction induced anti-inflammatory effects by reducing inflammation by >70 %, while antioxidant test revealed significant IC50 values for flavonoid content fraction (30.67 ± 2.09 µg/ml) and ethyl acetate fraction (42.66 ± 0.93 µg/ml). The maximum inhibition of acetylcholinesterase was exhibited by scopoletin content fraction (38.43 ± 3.94 %), while ethyl acetate fraction exerted neuroprotective effect against ß-amyloid peptide (83.97 ± 5.03 %). Phytochemical analysis, showed the presence of 3-O-quercetin glucopyranoside (415 mg/g), rutin (229.9 mg/g), ursolic and oleanolic acid (54 and 20.8 mg/g respectively), 3-O-quercetin rhamnopyranoside (12.8 mg/g), chlorogenic acid (9.5 mg/g), and scopoletin (1.38 mg/g). Our findings support the use of B. ternifolia since the extract induced significant neuroprotection against ß-amyloid peptide, anti-inflammatory, antioxidant and anti-acetylcholinesterase effects that could be attributed to its contents of polyphenols, coumarins, and triterpenes, and encourage further studies for development of this extract as therapeutic agent in treatment of Alzheimer's disease.

[Epigenetics, environment and asthma]. / Epigenética, medio ambiente y asma.

Asthma is a chronic inflammatory disease of the respiratory tract with a complex genetic background influenced by the exposition to a series of environmental factors. Genetic studies can only elucidate part of the heritability and susceptibility of asthma and even though several diseases have an evident genetic etiology, only a fraction of the genes involved in their pathogenicity have been identified. The epigenetic regulation of the latter is a fact one should bear in mind in order to explain the major triggers of diseases whose understanding is complicated, such as allergies and asthma. External stimulus such as nourishment, stress, physical activity, atmospheric pollution, tobacco smoking and alcohol drinking can induce either gene silencing or gene expression. In this regard, epigenetics can explain how these environmental factors influence our genetic inheritance. There is growing evidence that backs-up the fact that DNA methylation, histone post-translational modification and microRNA expression are influenced by the environment. This helps explaining how several of the risk factors mentioned contribute to the development and inheritance of asthma. In this review, different environmental factors and their relation with the main epigenetic regulatory mechanisms will be analyzed, as well as their possible role in the development of asthma.

El asma es una enfermedad inflamatoria crónica que afecta las vías respiratorias y tiene un componente genético complejo, que está mediada por la exposición a una variedad de desencadenantes ambientales. Los estudios genéticos no aclaran en su totalidad la herencia y susceptibilidad al asma. Muchas enfermedades son de origen genético; sin embargo, sólo se ha encontrado una fracción de los genes que las explican. La epigenética puede utilizarse para esclarecer las causas principales de padecimientos que son difíciles de entender, como la alergia y el asma. Estímulos externos como la nutrición, el estrés, la actividad física, la contaminación atmosférica y el consumo de tabaco y alcohol pueden silenciar o activar los genes. Al respecto, la epigenética ofrece explicaciones de cómo estos factores modifican sutilmente la herencia. Cada vez hay más evidencias que demuestran que los marcadores epigenéticos reconocidos, como la metilación del ADN, la modificación de histonas y la expresión de microRNAs están influidos por el ambiente. Esto ayuda a entender cómo muchos factores de riesgo similares a los señalados contribuyen a la aparición y herencia del asma. En esta revisión se analizan diferentes factores ambientales y su relación con los principales mecanismos epigenéticos, así como su posible influencia en la aparición del asma.

Monocyte locomotion inhibitory factor produced by E. histolytica improves motor recovery and develops neuroprotection after traumatic injury to the spinal cord.

Monocyte locomotion inhibitory factor (MLIF) is a pentapeptide produced by Entamoeba histolytica that has a potent anti-inflammatory effect. Either MLIF or phosphate buffered saline (PBS) was administered directly onto the spinal cord (SC) immediately after injury. Motor recovery was evaluated. We also analyzed neuroprotection by quantifying the number of surviving ventral horn motor neurons and the persistence of rubrospinal tract neurons. To evaluate the mechanism through which MLIF improved the outcome of SC injury, we quantified the expression of inducible nitric oxide synthase (iNOS), interleukin-10 (IL-10), and transforming growth factor- ß (TGF- ß ) genes at the site of injury. Finally, the levels of nitric oxide and of lipid peroxidation were also determined in peripheral blood. Results showed that MLIF improved the rate of motor recovery and this correlated with an increased survival of ventral horn and rubrospinal neurons. These beneficial effects were in turn associated with a reduction in iNOS gene products and a significant upregulation of IL-10 and TGF- ß expression. In the same way, MLIF reduced the concentration of nitric oxide and the levels of lipid peroxidation in systemic circulation. The present results demonstrate for the first time the neuroprotective effects endowed by MLIF after SC injury.

Therapeutic window for combination therapy of A91 peptide and glutathione allows delayed treatment after spinal cord injury.

Immunisation with neural-derived peptides is a promising strategy in models of spinal cord (SC) injury. Recent studies have also demonstrated that the addition of glutathione monoethyl ester (GHSE) to this strategy further improves motor recovery, tissue protection and neuronal survival after SC injury. As it is realistic to envision that this combination therapy could be tested in clinical trials, the therapeutic window should be experimentally explored before implementing its use in SC-injured human beings. For this purpose, 50 rats (10 per group) were subjected to a moderate SC contusion. The combined therapy was initiated at 10 min., 24, 72 or 120 hr after injury. Motor recovery and the survival of rubrospinal (RS) and ventral horn (VH) neurones were evaluated 60 days after injury. Results showed a significant motor improvement even if the combined therapy was initiated up to 72 hr after injury. BBB scores were as follows: 10 min.: 10.5 ± 0.7, 24 hr: 10.7 ± 0.5, 72 hr: 11.0 ± 1.3 and PBS: 6.7 ± 1 (mean ± S.D.). Initiation of combined therapy 120 hr after injury had no beneficial effect on motor recovery. Survival of RS and VH neurones was significantly higher in animals treated during the first 72 hr than those treated only with PBS. In this case again, animals treated with combined therapy 120 hr after injury did not present significant survival of neurones. Treatment with this combined strategy has a clinically feasible therapeutic window. This therapy provides enough time to transport and diagnose the patient and allows the concomitant use of other neuroprotective therapies.

Immunization with A91 peptide or copolymer-1 reduces the production of nitric oxide and inducible nitric oxide synthase gene expression after spinal cord injury.

Immunization with neurally derived peptides (INDP) boosts the action of an autoreactive immune response that has been shown to induce neuroprotection in several neurodegenerative diseases, especially after spinal cord (SC) injury. This strategy provides an environment that promotes neuronal survival and tissue preservation. The mechanisms by which this autoreactive response exerts its protective effects is not totally understood at the moment. A recent study showed that INDP reduces lipid peroxidation. Lipid peroxidation is a neurodegenerative phenomenon caused by the increased production of reactive nitrogen species such as nitric oxide (NO). It is possible that INDP could be interfering with NO production. To test this hypothesis, we examined the effect of INDP on the amount of NO produced by glial cells when cocultured with autoreactive T cells. We also evaluated the amount of NO and the expression of the inducible form of nitric oxide synthase (iNOS) at the injury site of SC-injured animals. The neural-derived peptides A91 and Cop-1 were used to immunize mice and rats with SC injury. In vitro studies showed that INDP significantly reduces the production of NO by glial cells. This observation was substantiated by in vivo experiments demonstrating that INDP decreases the amount of NO and iNOS gene expression at the site of injury. The present study provides substantial evidence on the inhibitory effect of INDP on NO production, helpingour understanding of the mechanisms through which protective autoimmunity promotes neuroprotection.

Anti-inflammatory defense mechanisms of Entamoeba histolytica.

The monocyte locomotion inhibitory factor (MLIF), a heat-stable oligopeptide found in the supernatant fluid of Entamoeba histolytica axenic cultures, may contribute to the delayed inflammation observed in amoebic hepatic abscess. This factor was isolated by ultra-filtration and high powered liquid chromatography, obtaining a primary Met-Gln-Cys-Asn-Ser structure, identified afterwards as the carboxyl-terminal (…Cys-Asn-Ser) active site. The selective anti-inflammatory effects of the pentapeptide have been observed in both in vitro and in vivo models, using a synthetic pentapeptide to maintain the same anti-inflammatory conditions during the experimental assays. Anti-inflammatory effects observed include inhibition of human monocyte locomotion and the respiratory burst in monocytes and neutrophils, increasing expression of anti-inflammatory cytokines and inhibiting expression of the adhesion molecules VLA-4 and VCAM, among others. In this review, we will describe the effects of MLIF detected so far and how it might be used as a therapeutical agent against inflammatory diseases.

Frequency of the serological reactivity against the caprine arthritis encephalitis lentivirus gp135 in children who consume goat milk.

BACKGROUND: Caprine arthritis encephalitis virus (CAEV) is a retrovirus belonging to the lentivirus genus that also includes the human immunodeficiency virus (HIV). CAEV may be transmitted to humans by goat milk consumption. It has been suggested that CAEV may also be involved in the immunological protection process against HIV, but this has not been demonstrated. Here we identified serological reactivity against CAEV gp135 in children who consumed goat milk. METHODS: Thirty sera samples from children (males between 6 and 16 years of age) who regularly consumed goat milk and a negative control of 30 serum samples from children (males between 6 and 12 years) with no previous contact with goats or goat dairy products were used. All sera were tested by Western blot against CAEV antigens. RESULTS: There were 18/30 serum samples from goat milk consumers that were reactive to CAEV gp135, and one reacted against gp50 simultaneously; none of the 30 serum samples from nonconsumers of goat dairy products reacted to viral proteins. CONCLUSIONS: These results showed that the positive response to gp135 may be the result of a repetitive stimulation without viral replication or the result of CAEV replication in humans. CAEV gp135 is codified by the env gene located on the viral particle surface as well as gp50. Moreover, there are similarities between CAEV gp135 and HIV-1 gp120, so there is a possibility that CAEV replicates in humans and may participate in immunological cross-phenomena, but this should be further studied.

The Cys-Asn-Ser carboxyl-terminal end group is the pharmacophore of the amebic anti-inflammatory monocyte locomotion inhibitory factor (MLIF).

The monocyte locomotion inhibitory factor (MLIF) is an anti-inflammatory oligopeptide produced by Entamoeba histolytica. Among its different effects, it inhibits locomotion of human monocytes, hence its original name. Previous experimental studies have shown that the anti-inflammatory properties of MLIF (Met-Gln-Cys-Asn-Ser) remained when aminoacid glutamine was substituted by a proline in the second position (pMLIF: Met-Pro-Cys-Asn-Ser). By changing the order of MLIF amino acids, the resulting scrambled oligopeptide (sMLIF: Gln-Cys-Met-Ser-Asn) has failed activity. By means of ab initio study at the Hartree-Fock and Density Functional Theory levels, it was found that MLIF and pMLIF peptides maintain a great structural similarity among the last three amino acids (...Cys-Asn-Ser) predicting a pharmacophore. The objective of this work was to experimentally verify in vivo and in vitro the existence of the pharmacophore group in MLIF. We assayed three tripeptides by respiratory burst and delayed hypersensitivity skin reactions. The tripeptide Cys-Asn-Ser carboxyl-terminal end group maintained 100% of its biological properties, as well as the anti-inflammatory activity of MLIF, while the other tripeptides tested did not do that.