Nitrate transporter protein NPF5.12 and major latex-like protein MLP6 are important defense factors against Verticillium longisporum.

Plant defense responses to the soil-borne fungus Verticillium longisporum causing stem stripe disease on oilseed rape (Brassica napus) are poorly understood. In this study, a population of recombinant inbred lines (RILs) using the Arabidopsis accessions Sei-0 and Can-0 was established. Composite interval mapping, transcriptome data, and T-DNA mutant screening identified the NITRATE/PEPTIDE TRANSPORTER FAMILY 5.12 (AtNPF5.12) gene as being associated with disease susceptibility in Can-0. Co-immunoprecipitation revealed interaction between AtNPF5.12 and the MAJOR LATEX PROTEIN family member AtMLP6, and fluorescence microscopy confirmed this interaction in the plasma membrane and endoplasmic reticulum. CRISPR/Cas9 technology was applied to mutate the NPF5.12 and MLP6 genes in B. napus. Elevated fungal growth in the npf5.12 mlp6 double mutant of both oilseed rape and Arabidopsis demonstrated the importance of these genes in defense against V. longisporum. Colonization of this fungus depends also on available nitrates in the host root. Accordingly, the negative effect of nitrate depletion on fungal growth was less pronounced in Atnpf5.12 plants with impaired nitrate transport. In addition, suberin staining revealed involvement of the NPF5.12 and MLP6 genes in suberin barrier formation. Together, these results demonstrate a dependency on multiple plant factors that leads to successful V. longisporum root infection.

Genome-Wide Characterization and Expression Analyses of Major Latex Protein Gene Family in Populus simonii × P. nigra.

Major latex proteins, or MLPs, are crucial to plants' capacity to grow, develop, and endure biotic and abiotic stresses. The MLP gene family has been found in numerous plants, but little is known about its role in Populus simonii × P. nigra. This study discovered and assessed 43 PtMLP genes that were unevenly dispersed throughout 12 chromosomes in terms of their physicochemical characteristics, gene structure, conserved motifs, and protein localization. Based on their phylogeny and protein structural characteristics, three separate subclasses of PtMLP family were identified. Segmental and tandem duplication were found to be essential variables in the expansion of the PtMLP genes. The involvement of the PtMLP genes in growth and development, as well as in the responses to different hormones and stresses, was demonstrated by cis-regulatory element prediction. The PtMLP genes showed varying expression patterns in various tissues and under different conditions (cold, salt, and drought stress), as demonstrated in RNA-Seq databases, suggesting that PsnMLP may have different functions. Following the further investigation of the genes demonstrating notable variations in expression before and after the application of three stresses, PsnMLP5 was identified as a candidate gene. Subsequent studies revealed that PsnMLP5 could be induced by ABA treatment. This study paves the way for further investigations into the MLP genes' functional mechanisms in response to abiotic stressors, as well as the ways in which they can be utilized in poplar breeding for improved stress tolerance.

Genome-wide characterization of major latex protein gene family in peanut and expression analyses under drought and waterlogging stress.

Peanut is an important oilseed crop around the world which provides vegetable oil, protein and vitamins for humans. Major latex-like proteins (MLPs) play important roles in plant growth and development, as well as responses to biotic and abiotic stresses. However, their biological function in peanut is still unclear. In this study, a genome-wide identification of MLP genes in cultivated peanut and two diploid ancestor species was analyzed to determine their molecular evolutionary characteristics and the expression profile under drought and waterlogging stress conditions. Firstly, a total of 135 MLP genes were identified from the genome of tetraploid peanut (Arachis hypogaea) and two diploid species Arachis. duranensis and Arachis. ipaensis. Then, phylogenetic analysis revealed that MLP proteins were divided into five different evolutionary groups. These genes were distributed unevenly at the ends of chromosomes 3, 5, 7, 8, 9 and 10 in three Arachis species. The evolution of MLP gene family in peanut was conserved and led by tandem and segmental duplication. The prediction analysis of cis-acting elements showed that the promoter region of peanut MLP genes contained different proportions of transcription factors, plant hormones-responsive elements and so on. The expression pattern analysis showed that they were differentially expressed under waterlogging and drought stress. These results of this study provide a foundation for further research on the function of the important MLP genes in peanut.

Overexpression of major latex protein 423 (NtMLP423) enhances the chilling stress tolerance in Nicotiana tabacum.

Chilling stress impedes plant growth and hinders crop development and productivity. In this study, we identified the major latex protein (MLP) in tobacco (NtMLP423) and examined its roles in chilling resistance. NtMLP423 expression was considerably upregulated in response to chilling stress. NtMLP423 function was assessed and compared in plants with overexpression and antisense characteristics. Under chilling stress, plants with overexpression characteristics grew better than wild-type and antisense plants. NtMLP423 overexpression reduced membrane lipid damage, increased antioxidant enzyme activity, and reduced reactive oxygen species (ROS) accumulation under chilling stress. Here, we screened for the first time the upstream transcription factor NtMYB108, which regulates NtMLP423 expression under chilling stress. The NtMYB108 transcription factor directly binds to the NtMLP423 promoter and improves NtMLP423 resistance to chilling stress. Subjecting NtMYB018 to virus-induced gene silencing reduced chilling stress tolerance. Overall, NtMLP423 overexpression enhances chilling stress tolerance, while its suppression has the opposite effect.

Combined Protein and Alkaloid Research of Chelidonium majus Latex Reveals CmMLP1 Accompanied by Alkaloids with Cytotoxic Potential to Human Cervical Carcinoma Cells.

Chelidonium majus L. is a latex-bearing plant used in traditional folk medicine to treat human papillomavirus (HPV)-caused warts, papillae, and condylomas. Its latex and extracts are rich in many low-molecular compounds and proteins, but there is little or no information on their potential interaction. We describe the isolation and identification of a novel major latex protein (CmMLP1) composed of 147 amino acids and present a model of its structure containing a conserved hydrophobic cavity with high affinity to berberine, 8-hydroxycheleritrine, and dihydroberberine. CmMLP1 and the accompanying three alkaloids were present in the eluted chromatographic fractions of latex. They decreased in vitro viability of human cervical cancer cells (HPV-negative and HPV-positive). We combined, for the first time, research on macromolecular and low-molecular-weight compounds of latex-bearing plants in contrast to other studies that investigated proteins and alkaloids separately. The observed interaction between latex protein and alkaloids may influence our knowledge on plant defense. The proposed toolbox may help in further understanding of plant disease resistance and in pharmacological research.

New and rapid strategies for the diagnosis of bovine paratuberculosis "in situ" using latex particles.

The chemical coupling of a protoplasmatic antigen from Mycobacterium avium subsp. paratubeculosis onto core-shell carboxylated particles was investigated with the aim of producing latex-protein complexes to be used in immunoagglutination assays capable of detecting bovine paratuberculosis disease. For this purpose, sensitizations were carried out using both colored and not colored carboxylated latexes as well as the protoplasmatic antigen at pH close to its isoelectric point to favor the antigenic protein to approach the particle surface. In all cases, higher fractions of proteins were chemically-bound to carboxyl groups on the surface of the particles. The assessment of the performance of the visual immunoagglutination assays consisted of evaluating 111 sera from healthy and infected bovines with Mycobacterium avium subsp. paratuberculosis. Complexes obtained from the colored latex allowed an acceptable visual discrimination between the studied positive and negative sera. Most of the positive samples showed strong to very strong agglutination and only a few samples reacted weakly, i.e. a sensitivity of 70%. The specificity of the assay, on the other hand, was 86%. Therefore, this rapid detection technique allows an easy and inexpensive identification of animals possibly infected with paratuberculosis "in situ" in the herds.

Major latex protein-like encoding genes contribute to Rhizoctonia solani defense responses in sugar beet.

Sugar beets are attacked by several pathogens that cause root damages. Rhizoctonia (Greek for "root killer") is one of them. Rhizoctonia root rot has become an increasing problem for sugar beet production and to decrease yield losses agronomical measures are adopted. Here, two partially resistant and two susceptible sugar beet genotypes were used for transcriptome analysis to discover new defense genes to this fungal disease, information to be implemented in molecular resistance breeding. Among 217 transcripts with increased expression at 2 days post-infection (dpi), three resistance-like genes were found. These genes were not significantly elevated at 5 dpi, a time point when increased expression of three Bet v I/Major latex protein (MLP) homologous genes BvMLP1, BvMLP2 and BvML3 was observed in the partially resistant genotypes. Quantitative RT-PCR analysis on diseased sugar beet seedlings validated the activity of BvMLP1 and BvMLP3 observed in the transcriptome during challenge by R. solani. The three BvMLP genes were cloned and overexpressed in Arabidopsis thaliana to further dissect their individual contribution. Transgenic plants were also compared to T-DNA mutants of orthologous MLP genes. Plants overexpressing BvMLP1 and BvMLP3 showed significantly less infection whereas additive effects were seen on Atmlp1/Atmlp3 double mutants. The data suggest that BvMLP1 and BvMLP3 may contribute to the reduction of the Rhizoctonia root rot disease in sugar beet. Impact on the defense reaction from other differential expressed genes observed in the study is discussed.

A lateral flow immunoassay based on colored latex particles for detection of canine visceral leishmaniasis.

Canine visceral leishmaniasis (CVL) is the major source of human visceral leishmaniasis. To control the spread of this disease, early and accurate detection of infected dogs is critical but challenging. The serological diagnosis of CVL remains problematic because there are no reliable commercially available tests. Most laboratories use enzyme-linked immunosorbent assay or the indirect immunofluorescent antibody test. These tests use Leishmania chagasi recombinant antigens K39 or K26 assembled with either gold-labelled Staphylococcus aureus protein A or protein G from Streptococcus pyogenes. In this work, we propose the development, optimization and standardization of a lateral flow immunoassay (LFIA) based on functionalized colored particles and a specific recombinant antigen, as a visual in situ method for the diagnosis of CVL. The following analysis variables were considered: (i) the concentration of the latex-protein complex; (ii) the dilution of the serum; (iii) the composition of the employed buffers; (iv) the nominal capillary flow time through the nitrocellulose membrane; (v) the concentration of reagents fixed in the test and control lines; (vi) the particle size of the colored latex; and (vii) the conjugation method. Then, the obtained strips were evaluated as a visual diagnostic tool based on a panel of positive and negative sera. It was observed that because of its simplicity and performance the LFIA test is a quick and reliable alternative for the diagnosis of CVL either in conventional laboratories or for remote areas where laboratories are not readily accessible for conventional assays.

Concentration-dependent effects of latex F1-protein fraction incorporated into deproteinized bovine bone and biphasic calcium phosphate on the repair of critical-size bone defects.

F1-protein fraction (F1) is a natural bioactive compound extracted from the rubber tree, Hevea brasiliensis, and has been recently studied for its therapeutic potential in wound healing. In this study, we investigated the concentration-dependent effects of F1 (0.01%, 0.025%, 0.05%, and 0.1%) incorporated into deproteinized bovine bone (DBB) and porous biphasic calcium phosphate (pBCP), on the repair of rat calvarial critical-size bone defects (CSBD). The defects were analyzed by 3D-microtomography and 2D-histomorphometry at 12 weeks postsurgery. The binding efficiency of F1 to pBCP (96.3 ± 1.4%) was higher than that to DBB (67.7 ± 3.3%). In vivo analysis showed a higher bone volume (BV) gain in all defects treated with DBB (except in 0.1% of F1) and pBCP (except in 0.05% and 0.1% of F1) compared to the CSBD without treatment/control group (9.96 ± 2.8 mm3 ). DBB plus 0.025% F1 promoted the highest BV gain (29.7 ± 2.2 mm3 , p < .0001) compared to DBB without F1 and DBB plus 0.01% and 0.1% of F1. In the pBCP group, incorporation of F1 did not promote bone gain when compared to pBCP without F1 (15.9 ± 4.2 mm3 , p > .05). Additionally, a small BV occurred in defects treated with pBCP plus 0.1% F1 (10.4 ± 1.4 mm3, p < .05). In conclusion, F1 showed a higher bone formation potential in combination with DBB than with pBCP, in a concentration-dependent manner. Incorporation of 0.25% F1 into DBB showed the best results with respect to bone formation/repair in CSBD. These results suggest that DBB plus 0.25% F1 can be used as a promising bioactive material for application in bone tissue engineering.

Genome-wide identification and expression analysis of major latex protein (MLP) family genes in the apple (Malus domestica Borkh.) genome.

Major latex protein/ripening-related proteins (MLP/RRP) subfamily are a class of proteins that play crucial roles in response to defense and stress response. However, their biological function is still not clear, the identification and characterization will provide essential information for understanding their roles. Here, we carried out a genome-wide evolutionary characteristics and gene expression analysis of the MLP family in apple (Malus domestica, Borkh.). A total of 36 MdMLP genes were screened in apple genome. They were uneven located on 5 chromosomes, where were mainly arranged in tandem clusters, and the phylogenetic analysis put forward further views on the evolutionary relationship and putative functions among the genes. The conserved motifs showed that the MLP proteins which contained motif 1 had the potential function, and tissue-specific expression analysis showed that apple MLP members had diverse biological roles. Furthermore, the results showed seven of the MdMLPs that harbored cis-acting regulatory elements in response to defense and stress, and our expression data proved that they were involved in biotic stresses. The present study provides new views to the evolution and regulation of MdMLP genes, which represent objectives of future research and incorporate in resistance-related molecular breeding projects.

Chemical constituents of Calotropis procera latex and ultrastructural effects on Haemonchus contortus / Constituintes químicos do látex de Calotropis procera e efeitos ultraestruturais em Haemonchus contortus

Abstract This study aimed to evaluate the anthelmintic and ultrastructural effects of Calotropis procera latex on Haemonchus contortus. C. procera latex was twice centrifuged at 10,000×g and dialyzed to obtain a fraction rich in proteins, named LP (latex protein), and at 3,000 rpm to obtain a fraction rich in secondary metabolites, named LNP (latex non-protein). Specimens of H. contortus exposed to LNP, LP and PBS in the Adult Worm Motility Test (AWMT) were submitted to scanning (SEM) and transmission (TEM) electron microscopy to verify changes in their ultrastructure. Phytochemical tests in the LNP indicated the presence of phenols, steroids, alkaloids and cardenolides. High-Performance Liquid Chromatography (HPLC) characterized the presence of the compounds gallic acid and quercetin in the LNP. The protein content in the LP was 43.1 ± 1.1 mg/mL and 7.7 ± 0.3 mg/mL in LNP. In AWMT, LNP and LP inhibited the motility of 100% of the nematodes, with LNP being more effective than LP and ivermectin more effective than both (p <0.05). Cuticle changes were observed by SEM and TEM in nematodes treated with LP and LNP. Calotropis procera latex has anthelmintic effects against H. contortus, causing damage to its cuticle and other alterations in its ultrastructure.

Resumo Este estudo objetivou avaliar os efeitos anti-helmínticos e ultraestruturais do látex de Calotropis procera sobre Haemonchus contortus. Látex de C. procera foi centrifugado duas vezes à a 10.000xg e dialisado para obter uma fração rica em proteínas, denominada proteínas do látex (LP). E centrifugado e centrifugado a 3.000 rpm, para obter uma fração rica em metabólitos secundários, denominada LNP (látex não proteico). Espécimes de H. contortus expostos à LNP, LP e PBS no Teste de Motilidade dos Nematoides Adultos (TMNA) foram submetidos a microscopia eletrônica de varredura (MEV) e de transmissão (MET), para verificar alterações em sua ultraestrutura. Testes fitoquímicos em LNP indicaram a presença de fenóis, esteroides, alcaloides e cardenolídeos. A presença dos compostos ácido gálico e quercetina em LNP foi caracterizada por Cromatografia Líquida de Alta Eficiência (CLAE). O conteúdo de proteínas em LP foi de 43,1 ± 1,1 mg/mL e de 7,7 ± 0,3 mg/mL em LNP. No TMNA, LNP e LP inibiram a motilidade de 100% dos nematoides, sendo LNP mais eficaz que LP, e a ivermectina mais eficaz que ambos (p <0,05). Alterações na cutícula de nematoides tratados com LP e LNP foram observadas por MEV e MET. O látex de C. procera apresenta efeito anti-helmíntico sobre H. contortus, causando danos à sua cutícula e outras alterações em sua ultraestrutura.

Genome-wide evolutionary characterization and expression analyses of major latex protein (MLP) family genes in Vitis vinifera.

The major latex protein/ripening-related protein (MLP/RRP) subfamily is known to be involved in a wide range of biological processes of plant development and various stress responses. However, the biological function of MLP/RRP proteins is still far from being clear and identification of them may provide important clues for understanding their roles. Here, we report a genome-wide evolutionary characterization and gene expression analysis of the MLP family in European Vitis species. A total of 14 members, was found in the grape genome, all of which are located on chromosome 1, where are predominantly arranged in tandem clusters. We have noticed, most surprisingly, promoter-sharing by several non-identical but highly similar gene members to a greater extent than expected by chance. Synteny analysis between the grape and Arabidopsis thaliana genomes suggested that 3 grape MLP genes arose before the divergence of the two species. Phylogenetic analysis provided further insights into the evolutionary relationship between the genes, as well as their putative functions, and tissue-specific expression analysis suggested distinct biological roles for different members. Our expression data suggested a couple of candidate genes involved in abiotic stresses and phytohormone responses. The present work provides new insight into the evolution and regulation of Vitis MLP genes, which represent targets for future studies and inclusion in tolerance-related molecular breeding programs.

Development of a simple and economical diagnostic test for canine leishmaniasis.

Visceral leishmaniasis is a public health problem worldwide. The early diagnosis in dogs is crucial, since they are an epidemiologically relevant reservoir of the disease. The aim of a field study is to early identify the disease allowing rapid intervention to reduce its effects. We propose an immunoagglutination test as a visual in situ method for diagnosis of canine visceral leishmaniasis. Latex-protein complexes were sensitized by covalent coupling of a chimeric recombinant antigen of Leishmania spp. onto polystyrene latex with carboxyl functionality. The reaction time and the antigen concentration under which the immunoagglutination assay shows greater discrimination between the responses of a positive control serum and a negative control serum were determined. Then, the latex-protein complexes were evaluated as a visual diagnostic tool with a panel of 170 sera. The test may be read between 2 and 5 min and can be performed even using sera with elevated concentration of lipids, bilirubin or with variable percentage of hemolysis. The sensitivity, the specificity and the diagnostic accuracy were 78%; 100% and >80%, respectively. The visual immunoagglutination test is of potential application as a method for field studies because it shows results in less than 5 min, it is easy to implement and does not require sophisticated equipment.

Diagnosis of toxoplasmosis in pregnancy. Evaluation of latex-protein complexes by immnunoagglutination.

The aim of this work was to obtain a reagent based on latex particles for ruling out acute toxoplasmosis in pregnant women by immunoagglutination (IA). Latex-protein complexes (LPC) were previously synthesized coupling the recombinant protein of Toxoplasma gondii P22Ag and the homogenate of the parasite to latex particles with different size, chemical functionality and charge density. LPC were tested in IA assays against a panel of 72 pregnant women serum samples. Results were analysed through receiver operating characteristic curves, determining area under the curve (AUC), sensitivity, specificity positive and negative predictive values (PPV and NPV, respectively). It was observed that the antigenicity of proteins was not affected during sensitization by either physical adsorption or covalent coupling. The best results in the sense of maximizing discrimination of low avidity sera from chronic ones were observed for the IA test based on latex particles with carboxyl functionality and the recombinant P22Ag, obtaining an AUC of 0·94, a sensitivity of 100% and a NPV of 100%. In this way, the proposed test could be useful for the toxoplasmosis diagnosis in pregnant women, with the advantages of being cheap, rapid and easy to be implemented.

Proteomic comparison of Chelidonium majus L. latex in different phases of plant development.

Chelidonium majus L. (Papaveraceae) latex is used in traditinonal folk medicine to treat papillae, warts, condylomas, which are visible effects of human papilloma virus (HPV) infections. The aim of this work was to provide new insights into the biology and medicinal use of C. majus milky sap in the flowering and fruit ripening period of the plant by comparing the protein content between samples collected on respective developmental stages using LC-MS-based label-free proteome approach. For quantification, the multiplexed LC-MS data were processed using comparative chemometric approach. Progenesis LC-MS results showed that in green fruit phase (stage IV), comparing to flowering phase (stage III) of plant development, a range of proteins with higher abundance were identified as stress- and defense-related. On the other hand at stage III very intense protein synthesis, processes of transcription, protein folding and active transport of molecules (ABC transporters) are well represented. 2-DE protein maps showed an abundant set of spots with similar MWs (about 30-35 kDa) and pIs (ca. 5.5-6.5), which were identified as major latex proteins (MLPs). Therefore we suggest that biological activity of C. majus latex could be related to its protein content, which shifts during plant development from intense biosynthetic processes (biosynthesis and transport of small molecules, like alkaloids) to plant defense mechanisms against pathogens. Further studies will help to elucidate if these defense-related and pathogenesis-related proteins, like MLP, together with small-molecule compounds, could inhibit viral infection, what could be a step to fully understand the medicinal activity of C. majus latex.

Combination of transcriptomic and proteomic approaches helps to unravel the protein composition of Chelidonium majus L. milky sap.

MAIN CONCLUSION: A novel annotated Chelidonium majus L. transcriptome database composed of 23,004 unique coding sequences allowed to significantly improve the sensitivity of proteomic C. majus assessments, which showed novel defense-related proteins characteristic to its latex. To date, the composition of Chelidonium majus L. milky sap and biosynthesis of its components are poorly characterized. We, therefore, performed de novo sequencing and assembly of C. majus transcriptome using Illumina technology. Approximately, 119 Mb of raw sequence data was obtained. Assembly resulted in 107,088 contigs, with N50 of 1913 bp and N90 of 450 bp. Among 34,965 unique coding sequences (CDS), 23,004 obtained CDS database served as a basis for further proteomic analyses. The database was then used for the identification of proteins from C. majus milky sap, and whole plant extracts analyzed using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) approach. Of about 334 different putative proteins were identified in C. majus milky sap and 1155 in C. majus whole plant extract. The quantitative comparative analysis confirmed that C. majus latex contains proteins connected with response to stress conditions and generation of precursor metabolites and energy. Notable proteins characteristic to latex include major latex protein (MLP, presumably belonging to Bet v1-like superfamily), polyphenol oxidase (PPO, which could be responsible for browning of the sap after exposure to air), and enzymes responsible for anthocyanidin, phenylpropanoid, and alkaloid biosynthesis.

Cotton major latex protein 28 functions as a positive regulator of the ethylene responsive factor 6 in defense against Verticillium dahliae.

In this study, we identified a defense-related major latex protein (MLP) from upland cotton (designated GhMLP28) and investigated its functional mechanism. GhMLP28 transcripts were ubiquitously present in cotton plants, with higher accumulation in the root. Expression of the GhMLP28 gene was induced by Verticillium dahliae inoculation and was responsive to defense signaling molecules, including ethylene, jasmonic acid, and salicylic acid. Knockdown of GhMLP28 expression by virus-induced gene silencing resulted in increased susceptibility of cotton plants to V. dahliae infection, while ectopic overexpression of GhMLP28 in tobacco improved the disease tolerance of the transgenic plants. Further analysis revealed that GhMLP28 interacted with cotton ethylene response factor 6 (GhERF6) and facilitated the binding of GhERF6 to GCC-box element. Transient expression assay demonstrated that GhMLP28 enhanced the transcription factor activity of GhERF6, which led to the augmented expression of some GCC-box genes. GhMLP28 proteins were located in both the nucleus and cytoplasm and their nuclear distribution was dependent on the presence of GhERF6. Collectively, these results demonstrate that GhMLP28 acts as a positive regulator of GhERF6, and synergetic actions of the two proteins may contribute substantially to protection against V. dahliae infection in cotton plants.

Properties of the tibialis anterior muscle after treatment with laser therapy and natural latex protein following sciatic nerve crush.

INTRODUCTION: In this study we evaluated the characteristics of the tibialis anterior muscle after sciatic nerve crush and treatment with low-level laser therapy (LLLT) or the protein from natural latex (P1). METHODS: We studied the following 6 groups of male Wistar rats: control (CG); exposed nerve (EG); injured nerve (IG); injured nerve with LLLT (LG); injured nerve with P1 (PG); and injured nerve with P1 and LLLT (LPG). RESULTS: After 4 weeks, muscle morphology showed improvement in the treated groups; after 8 weeks, the treated groups resembled controls, especially the PG. Morphometry revealed muscle fiber atrophy after nerve injury, with time-dependent recovery. Histochemical analysis revealed increased intermediate fiber area. The PG was more similar to controls with NADH staining, whereas the LPG more closely resembled controls with SDH staining. CONCLUSION: Treatment using only P1 proved most efficient, revealing a negative interaction between P1 and LLLT.

Morphometric and high resolution scanning electron microscopy analysis of low-level laser therapy and latex protein (Hevea brasiliensis) administration following a crush injury of the sciatic nerve in rats.

This study evaluated the effect of low-level laser therapy (LLLT; 15 J/cm(2)) and a latex protein (F1) on a crush injury of the sciatic (ischiadicus) nerve. Seventy-two rats (male, 250 g) were divided into 6 groups: CG, control; EG, exposed nerve; IG, injured nerve without treatment; LG, injured nerve with LLLT; HG, injured nerve with F1; and LHG, injured nerve with LLLT and F1. After 4 or 8 weeks, the animals were euthanized and samples of the sciatic nerve were collected for morphometric and high-resolution scanning electron microscopy (HRSEM) analysis. After 4 weeks, the morphometry revealed improvements in the treated animals, and the HG appeared to be the most similar to the CG; after 8 weeks, the injured groups showed improvements compared to the previous period, and the results of the treatment groups were more similar to one another. At HRSEM after 4 weeks, the treated groups were similar and showed improvement compared to the IG; after 8 weeks, the LHG and HG had the best results. In conclusion, the treatments resulted in improvement after the nerve injury, and this recovery was time-dependent. In addition, the use of the F1 resulted in the best morphometric and ultrastructural findings.

Morphological and morphometric analyses of crushed sciatic nerves after application of a purified protein from natural latex and hyaluronic acid hydrogel.

Hyaluronic acid hydrogels (HAHs) have been used as a carrier of substances and factors in the repair of nervous tissue. Natural latex protein (Hevea brasiliensis, F1) has shown positive effects in treating various types of tissues, including peripheral nerves. This study evaluated the F1 associated with a HAH in a controlled crush injury (axonotmesis) of the sciatic nerve in Wistar rats. The samples were photomicrographed for morphometric and quantitative analyzes using ImageJ 1.47k software (NIH, Bethesda, MD). Morphological, quantitative (myelin area/nerve area ratio and capillary density) and morphometric (minimum nerve fiber diameter, G-Ratio) data revealed an improvement in the recovery of the sciatic nerve with the application of HAH and the combination of HAH and F1 after 4 and 8 weeks of nerve injury. The most efficacious results were observed with the combination of both substances, F1 and HAH, revealing the regenerative capacity of this new biomaterial, which was hardly tested on nerve tissue.