Telocytes: current methods of research, challenges and future perspectives.

Telocytes (TCs) are CD34-positive interstitial cells that have long cytoplasmic projections, called telopodes; they have been identified in several organs and in various species. These cells establish a complex communication network between different stromal and epithelial cell types, and there is growing evidence that they play a key role in physiology and pathology. In many tissues, TC network impairment has been implicated in the onset and progression of pathological conditions, which makes the study of TCs of great interest for the development of novel therapies. In this review, we summarise the main methods involved in the characterisation of these cells as well as their inherent difficulties and then discuss the functional assays that are used to uncover the role of TCs in normal and pathological conditions, from the most traditional to the most recent. Furthermore, we provide future perspectives in the study of TCs, especially regarding the establishment of more precise markers, commercial lineages and means for drug delivery and genetic editing that directly target TCs.

Ultrastructural and Molecular Development of the Myotendinous Junction Triggered by Stretching Prior to Resistance Exercise.

The myotendinous junction (MTJ) is a highly specialized region of the locomotor apparatus. Here, we investigated the ultrastructural and molecular effects in the MTJ region after static stretching prior to the ladder-based resistance training. Thirty-two male, 60-day old Wistar rats were divided into four groups: Sedentary, Resistance Training, Stretching, and Stretching-Resistance Training. The gastrocnemius muscle was processed for transmission electron microscopy techniques and Western blot assay. We observed that the static stretching prior to the ladder-based resistance training increased the MTJ components, the fibroblast growth factor (FGF)-2 and FGF-6 protein expression. Also, we demonstrated the lower transforming growth factor expression and no difference in the lysyl oxidase expression after combined training. The MTJ alterations in response to combined training demonstrate adaptive mechanisms which can be used for the prescription or development of methods to reduce or prevent injuries in humans and promote the myotendinous interface benefit.

Structural and Ultrastructural Changes in the Tongue of mdx Mice.

The mdx mouse is an experimental model of Duchenne muscular dystrophy, a genetic disorder characterized by progressive muscular degeneration which affects the oral cavity musculature, and promotes difficulty in swallowing. This study aimed to describe morphological, structural, and ultrastructural changes in the tongue mucosa and musculature of mdx mice. Forty six-month-old mice were divided into two groups: Control C57bl/10 (n = 20) and mdx C57bl/10mdx (n = 20). The tongue was dissected and analyzed with light microscopy, scanning electron microscopy, and transmission electron microscopy techniques. Our results showed conical and triangular filiform, fungiform, foliate, and vallate papillae, and their connective tissue cores. The epithelium layers identified were corneum, granulosum, spinosum, and basale. The mdx group had a thicker epithelium. Lamina propria was composed of reddish and greenish collagen. In mdx, collagen was present in the musculature of the tongue's body and in the muscular tissue between mucous and serous glands of the caudal region. Musculature was also characterized by a shorter length of sarcoplasmic invaginations, myocytolysis in mitochondrial groupings, and inflammatory focus. In conclusion, the tongue of 6-month-old mdx mice had morphology, structure, and ultrastructure revealed, showing higher wear of filiform papillae indirect reflex from the muscular degeneration process.

Crotalphine Attenuates Pain and Neuroinflammation Induced by Experimental Autoimmune Encephalomyelitis in Mice.

Multiple sclerosis (MS) is a demyelinating disease of inflammatory and autoimmune origin, which induces sensory and progressive motor impairments, including pain. Cells of the immune system actively participate in the pathogenesis and progression of MS by inducing neuroinflammation, tissue damage, and demyelination. Crotalphine (CRO), a structural analogue to a peptide firstly identified in Crotalus durissus terrificus snake venom, induces analgesia by endogenous opioid release and type 2 cannabinoid receptor (CB2) activation. Since CB2 activation downregulates neuroinflammation and ameliorates symptoms in mice models of MS, it was presently investigated whether CRO has a beneficial effect in the experimental autoimmune encephalomyelitis (EAE). CRO was administered on the 5th day after immunization, in a single dose, or five doses starting at the peak of disease. CRO partially reverted EAE-induced mechanical hyperalgesia and decreased the severity of the clinical signs. In addition, CRO decreases the inflammatory infiltrate and glial cells activation followed by TNF-α and IL-17 downregulation in the spinal cord. Peripherally, CRO recovers the EAE-induced impairment in myelin thickness in the sciatic nerve. Therefore, CRO interferes with central and peripheral neuroinflammation, opening perspectives to MS control.

Morphological Changes in the Motor Endplate and in the Belly Muscle Induced by Previous Static Stretching to the Climbing Protocol.

Static stretching provides benefits to the range of motion, modulates intramuscular connective tissue, and is incorporated into warm-up exercises. In this study, we present the effects in the motor endplate and belly muscle resulting from previous static stretching to climbing training. Twenty-four adult male Wistar rats were divided into four groups (n = 6 each): Sedentary (Sed), Climbing (Clb), Static stretching (Ss), and Static stretching prior to climbing (Ssc). The animals (Clb, Ss, and Ssc groups) were subjected to a training protocol 3×/week for 8 weeks, and the Ssc group was subjected to the Ss and Clb protocols in the same session. Samples from the animals were processed for immunostaining, histochemistry, and light microscopy. The Clb group presented a higher motor endplate; the Ss group presented no changes in the motor endplate; and the Ssc group demonstrated a higher compactness. We concluded that static stretching prior to the climbing protocol maintained the density of the motor endplate and increased the compactness of the neuromuscular junction structure. Also, there was a reduction in the myofibers' diameter (Type I and IIa), an increase in myofibrillar densities (Type I and IIx, and total), and the reorganization of the myonuclei and the interstitium.

Myotendinous Junction Components of Different Skeletal Muscles Present Morphological Changes in Obese Rats.

Obesity is characterized by excess adipose tissue and chronic inflammation and promotes extensive changes that can compromise skeletal muscles' structural and functional integrity. Obesity can seriously impact the force transmission region between the muscle and the tendon, the myotendinous junction (MTJ). The present study aimed to investigate the plasticity of muscle fibers and MTJ regions in high-fat diet-induced obesity in rat tibialis anterior (TA) and soleus (SO) muscles. Wistar rats were divided into control and obese groups (induced by a high-fat diet). The samples of TA and SO muscles were prepared for histochemical and ultrastructural analysis (sarcomeres and MTJ projection). In the muscle fiber, similar adaptations were observed between the muscles of the smaller fiber (types I and IIa) in the obesity results. The MTJ region demonstrated different adaptations between the analyzed muscles. The TA­MTJ region has shorter ultrastructures, while in the SO­MTJ region, the ultrastructures were larger. We conclude that obesity induced by a high-fat diet promotes similar adaptation in the muscle fibers; however, in the MTJ region, the sarcoplasmatic projections and adjacent sarcomere demonstrate different adaptations according to distinct muscle phenotypes.

Structural and ultrastructural characterization of the palatine epithelium of the Guinea pig: A new record of telocytes in the oral cavity.

The morphology of the oral cavity of mammals relates to diet, habitat, and function. The palate is an important region with adaptations for oral somatosensation and mechanical loads due to the pressure of the tongue with food. The research aimed to describe the structural and ultrastructural characteristics of the epithelium and the connective tissue cores of the guinea pig palate using macroscopic, light microscopy, scanning electron microscopy, and transmission electron microscopy analysis. The hard palate had conical and filiform papillae, and the soft palate had open salivary ducts. After the removal of the epithelium, the connective tissue cores revealed thin filaments and laminar projections in the hard palate, and opening ducts were evidenced in the soft palate. The palatine epithelium was keratinized and organized by layers, lamellated corpuscles were found in lamina propria of the hard palate. In contrast, the soft palate had glands clusters associated with nerve fibers, and in both regions were identified telocytes. We concluded that the hard palate presented conical and filiform papillae that differ from other mammals. Besides, it is a new description of the connective tissue cores morphology and the first record of the telocytes in this anatomical region for mammals.

Crotalphine attenuates pain and neuroinflammation induced by experimental autoimmune encephalomyelitis in mice

Multiple sclerosis (MS) is a demyelinating disease of inflammatory and autoimmune origin, which induces sensory and progressive motor impairments, including pain. Cells of the immune system actively participate in the pathogenesis and progression of MS by inducing neuroinflammation, tissue damage, and demyelination. Crotalphine (CRO), a structural analogue to a peptide firstly identified in Crotalus durissus terrificus snake venom, induces analgesia by endogenous opioid release and type 2 cannabinoid receptor (CB2) activation. Since CB2 activation downregulates neuroinflammation and ameliorates symptoms in mice models of MS, it was presently investigated whether CRO has a beneficial effect in the experimental autoimmune encephalomyelitis (EAE). CRO was administered on the 5th day after immunization, in a single dose, or five doses starting at the peak of disease. CRO partially reverted EAE-induced mechanical hyperalgesia and decreased the severity of the clinical signs. In addition, CRO decreases the inflammatory infiltrate and glial cells activation followed by TNF-α and IL-17 downregulation in the spinal cord. Peripherally, CRO recovers the EAE-induced impairment in myelin thickness in the sciatic nerve. Therefore, CRO interferes with central and peripheral neuroinflammation, opening perspectives to MS control.

Effects of Strength Training and Anabolic Steroid in the Peripheral Nerve and Skeletal Muscle Morphology of Aged Rats.

Thirty male 20-month-old Wistar rats were divided into groups: IC-initial control (n = 6), FC-final control (n = 6), AC-anabolic hormone control (n = 6), ST-strength trained (n = 6) and STA-strength trained with anabolic hormone (n = 6). All groups were submitted to adaptation, familiarization and maximum load carrying test (MLCT). Strength training (6-8×/session with loads of 50%-100% MLCT, 3×/week and pause of 120 s) was performed in ladder climbing (LC) for 15 weeks. The administration of testosterone propionate (TP) was performed 2×/week (10 mg/kg) in animals in the AC and STA groups. After the experimental period, animals were euthanized and the tibial nerve and plantaris muscle removed and prepared for electron transmission and histochemistry. To compare the groups we used one-way ANOVA (post hoc Bonferroni), student's t-tests for pre vs. post (dependent and independent variables) comparisons and significance level set at p ≤ 0.05. The following significant results were found: (a) aging decreased the number of myelinated axon fibers; (b) use of isolated TP increased the diameter of myelinated fibers, along with increased thickness of myelin sheath; (c) ST increased area of myelinated and unmyelinated fibers, together with the myelin sheath. These changes made it possible to increase the area occupied by myelinated fibers keeping their quantity and also reduce the interstitial space; and (d) association of anabolic steroid and ST increased the area of unmyelinated axons and thickness of the myelin sheath. Compared to ST, both strategies have similar results. However, Schwann cells increased significantly only in this strategy.

A preliminary investigation into the morphology of oral papillae and denticles of blue sharks (Prionace glauca) with inferences about its functional significance across life stages.

Sensory organs in elasmobranchs (sharks, skates, rays) detect and respond to a different set of biotic and/or abiotic stimuli, through sight, smell, taste, hearing, mechanoreception and electroreception. Although gustation is crucial for survival and essential for growth, mobility, and maintenance of neural activity and the proper functioning of the immune system, comparatively little is known about this sensory system in elasmobranchs. Here we present a preliminary investigation into the structural and dimensional characteristics of the oral papillae and denticles found in the oropharyngeal cavity of the blue shark (Prionace glauca) during embryonic development through adulthood. Samples were obtained from the dorsal and ventral surface of the oropharyngeal cavity collected from embryos at different development stages as well as from adults. Our results suggest that development of papillae occurs early in ontogeny, before the formation of the oral denticles. The diameter of oral papillae gradually increases during development, starting from 25 µm in stage I embryos, to 110 µm in stage IV embryos and 272-300 µm in adults. Embryos exhibit papillae at early developmental stages, suggesting that these structures may be important during early in life. The highest density of papillae was observed in the maxillary and mandibular valve regions, possibly related to the ability to identify, capture and process prey. The oral denticles were observed only in the final embryonic stage as well as in adults. Accordingly, we suggest that oral denticles likely aid in ram ventilation (through reducing the hydrodynamic drag), to protect papillae from injury during prey consumption and assist in the retention and consumption of prey (through adhesion), since these processes are only necessary after birth.

Aging Induces Changes in the Somatic Nerve and Postsynaptic Component without Any Alterations in Skeletal Muscles Morphology and Capacity to Carry Load of Wistar Rats.

The present study aimed to analyze the morphology of the peripheral nerve, postsynaptic compartment, skeletal muscles and weight-bearing capacity of Wistar rats at specific ages. Twenty rats were divided into groups: 10 months-old (ADULT) and 24 months-old (OLD). After euthanasia, we prepared and analyzed the tibial nerve using transmission electron microscopy and the soleus and plantaris muscles for cytofluorescence and histochemistry. For the comparison of the results between groups we used dependent and independent Student's t-test with level of significance set at p ≤ 0.05. For the tibial nerve, the OLD group presented the following alterations compared to the ADULT group: larger area and diameter of both myelinated fibers and axons, smaller area occupied by myelinated and unmyelinated axons, lower numerical density of myelinated fibers, and fewer myelinated fibers with normal morphology. Both aged soleus and plantaris end-plate showed greater total perimeter, stained perimeter, total area and stained area compared to ADULT group (p < 0.05). Yet, aged soleus end-plate presented greater dispersion than ADULT samples (p < 0.05). For the morphology of soleus and plantaris muscles, density of the interstitial volume was greater in the OLD group (p < 0.05). No statistical difference was found between groups in the weight-bearing tests. The results of the present study demonstrated that the aging process induces changes in the peripheral nerve and postsynaptic compartment without any change in skeletal muscles and ability to carry load in Wistar rats.

Morphological analysis of the oviduct, oviducal gland and isthmus of the blue shark Prionace glauca (Linnaeus, 1758) (Elasmobranchii: Carcharhiniformes)

Oviducal gland present in elasmobranchs is correlated to the organism's reproductive strategy, and its functions are to produce mucus, to form the egg's tertiary envelope and to store sperm. The gland contains four zones: club, papillary,baffle and terminal. The structures of the oviduct, oviducal gland and isthmus of blue shark Prionace glauca were described using macroscopic, light microscopy and scanning electron microscopy techniques. The epithelium of the oviduct and isthmus is folded and is a simple, columnar, ciliated lining epithelium with glandular cells. In the oviducal gland, the lining tissues in the four zones are similar to the oviduct and isthmus lining. The terminal zone shows the presence of sperm in the lumen of the secretory tubules, which remains stored even in the absence of recent copulation. Here, these organs were studied and their connections in an attempt to elucidate the mechanisms of reproduction in the blue shark, showing the three-dimensional aspects, thus adding morphological information important for the understanding of the structure and functioning of these organs of fundamental importance in the life of the majority of elasmobranchs.

A glândula oviducal presente nos elasmobrânquios está correlacionada com a estratégia reprodutiva, cuja função é de produzir o muco, formar o envelope terciário do ovo e armazenar espermatozóides. A glândula contém quatro zonas: club, papilar, baffle e terminal. Foi descrita a estrutura do oviduto, glândula oviducal e istmo do tubarão-azul, Prionace glauca , pelas técnicas macroscópica, microscópica de luz e eletrônica de varredura. Foi observado que no oviduto e istmo o epitélio é pregueado de revestimento simples, colunar, ciliado com células glandulares. Na glândula oviducal o tecido de revestimento é semelhante ao oviduto e istmo nas quatro zonas. Na zona terminal observou-se a presença de espermatozoides no lúmen dos túbulos secretores que permanecem estocados mesmo na ausência de copula recente. Aqui, estudaram-se esses órgãos e suas conexões na tentativa de elucidar os mecanismos da reprodução no tubarão azul, apresentando os aspectos tridimensionais, desta forma agregando informações morfológicas importantes para o entendimento da estrutura e funcionamento desses órgãos de fundamental importância na vida da maioria dos elasmobrânquios.

Neural mobilization and static stretching in an experimental sciatica model: an experimental study / Mobilização neural e alongamento estático em um modelo experimental de ciatalgia: estudo experimental

OBJECTIVE: To verify the effectiveness of neural mobilization and static stretching in reducing pain in rats submitted to experimental sciatica. METHODS: The rats (n=23) were divided into three groups: sham (SG/n=8), without intervention; stretching (STCG/n=8), treated with static stretching; and neural mobilization (NMG/n=7), treated with neural mobilization. The animals underwent an experimental model of sciatica by compression of the right ischiatic nerve with catgut suture thread. There were five consecutive sessions of treatment that began on the third day after lesion. The pain caused by the sciatica was evaluated by a functional incapacitation test that measured paw elevation time (PET), and values over 10s were indicative of pain. PET was measured at the following moments: before the lesion (M1), immediately before (M2) and after the first session (M3), immediately after the last session (M4) and 24h after the last session (M5). ANOVA was applied with repeated measures and unrepeated measures for intra- and inter-group comparison, respectively. RESULTS: In the SG, post-lesion PETs were greater than M1 (p<0.001), suggesting persistence of pain. In the STCG, post-lesion PETs were greater than M1 (p<0.001), but lower when comparing M3 vs. M4 (p<0.05) and M3 vs. M5 (p<0.01) suggesting the effectiveness of the treatment. In NMG, M2, M3 (p<0.001) and M4 (p<0.05) were greater in relation to M1, but not M5, showing that this treatment reestablished the normal PET values. CONCLUSION: Both forms of therapy were effective in reducing pain, with neural mobilization being the more effective of the two.

OBJETIVO: Verificar a eficácia da mobilização neural e do alongamento estático na redução da dor em ratos submetidos à ciática experimental. MÉTODOS: Os ratos (n=23) foram divididos em três grupos: simulacro (GS/n=8), sem intervenção; alongamento (GAL/n=8), tratados com alongamento estático; e mobilização neural (GMN/n=7), tratados com mobilização neural. Submeteram-se os animais a um modelo experimental de ciática, comprimindo o nervo isquiático direito com fio de catgut. Realizaram-se cinco sessões consecutivas de tratamento que se iniciaram no terceiro dia pós-lesão. Avaliou-se a dor, provocada pela ciática, pelo teste de incapacidade funcional que mensurava o tempo de elevação da pata (TEP) do animal, e valores maiores que 10 segundos eram indicativos de dor. O TEP foi mensurado nos momentos: antes da lesão (M1), imediatamente antes (M2), após a 1ª sessão (M3), imediatamente após a última sessão (M4) e 24 horas após a última sessão (M5). Aplicou-se ANOVA com medidas repetidas e não repetidas para análise intra e intergrupos, respectivamente. RESULTADOS: No GS, os TEPs no pós-lesão foram maiores que M1(p<0,001), sugerindo persistência da dor. No GAL, os TEPs foram maiores no pós-lesão em relação a M1 (p<0,001), mas diminuiu nas comparações M3xM4 (p<0,05) e M3xM5 (p<0,01) sugerindo a eficácia do tratamento. No GMN, M2, M3 (p<0,001) e M4 (p<0,05) foram maiores em relação a M1, mas M5 não, mostrando que este tratamento restabeleceu os valores normais de TEP. CONCLUSÃO: As duas formas de terapia foram eficazes na redução da dor, sendo a mobilização neural mais efetiva.