Bioindicators of the nanoscale structural quality of bone were investigated using ion milling and transmission electron microscopy of osteoporotic bone from human female donors. / Bone is a complex hierarchical biomaterial constantly undergoing remodeling events initiated by cell signaling and fulfilled by migratory bone cells. In osteoporosis, a multitude of signaling factors cause bone resorption to proceed quicker than bone reformation, resulting in a lower bone mineral density (BMD) and porosity as seen by thinning of the cortex and trabeculae. However, the structural motifs of these altered regions of the skeleton have not been understood on the nanoscale. In this thesis, transmission electron microscopy (TEM) was used with an image analysis technique termed nanomorphometry, developed to enable the measurement of nanoscale structural features in human bone. Several nanoscale bone quality bioindicators relevant to the collagen fibrils and bone mineral (mineral lamellae, ML) components were defined and tested (collagen fibril diameter, interfibrillar spacing, ML thickness & ML stack thickness) among two donor cohorts of post-menopausal osteoporotic female patients and age- and sex-matched controls. In one cohort, the anatomical region investigated was the intertrochanteric crest of the femur, while in the second, the femoral neck was studied. The bone sections were prepared using an ion milling workflow yielding electron-transparent views of the bone ultrastructure. Blinded image analysis of the ultrastructure revealed that in both cohorts, the thickness of the MLs was significantly larger in osteoporotic samples versus their controls. In the former cohort, it was found that anti-resorptive drug use in the treated group did not return the ML thickness back to control levels. In the latter cohort, the ML thickness correlated more closely with the proximal femur bone mineral density (BMD) than the age of the patient. These findings suggest that the morphology of the nanoscale mineral phase is affected by osteoporosis, an effect indirectly observed by other techniques, and warrants further exploration into the implications of this effect on bone quality, fragility and strength. / Thesis / Master of Applied Science (MASc) / Human bone is a biomaterial with many levels of organization from the macroscale down to the nanoscale. The material consists of roughly 30 weight % organic components (collagen, non-collagenous proteins) and 67 weight % inorganic components (calcium phosphate minerals) deposited by bone cells. Osteoporosis is a bone disease commonly associated with increased bone porosity and bone fragility. In this study, the effect of osteoporosis on the nanoscale structure of bone was directly imaged and investigated using transmission electron microscopy (TEM). Two advanced ion milling techniques (broad beam and focused ion beam) were used to thin the bone specimens for TEM. Bioindicators relating to the structure and size of collagen and mineral components in osteoporotic versus control bone were quantified in an unbiased image analysis workflow. Findings indicated an increase in the thickness of poly-crystalline bone mineral lamellae in the nanoscale structure of human osteoporotic bone from two human donor cohorts.

This thesis investigates the diversity of two silica-scaled protist groups, Paraphysomonadida and Thaumatomonadida by light and electron microscopical observations and sequencing (rDNA) on novel clonal cultures. Despite these groups of protist dominating pelagic, littoral as well as inland freshwater and soil habitats, they are taxonomically poorly understood to the extent that any progress in ecological theory is hampered. Now that environmental DNA sequencing is being carried out faster than we can characterise protists from culture it is important that we understand how molecular and physical diversity match up, especially because so many protists are morphospecies. Nearly one hundred isolates were cultured on which both morphological and molecular data was carried out in parallel to reveal around 50 new species of protist from eight different genera: two heterokont genera, Paraphysomonas and Incisomonas n. gen., and six cercozoan genera, Thaumatomonas, Allas, Reckertia, Thaumatospina n. gen., Cowlomonas n. gen., and Scutellomonas n. gen. These data make major contributions to taxonomy and understanding aspects of protist diversity where previously morphological diversity was heavily biased towards over- generalized morphotypes. This thesis quickly showed that gross lumping of morphospecies was true of Paraphysomonas, for which many of the isolates cultured herein might have been regarded as one species (not more than 20). The many cultured isolates exhibited varied cell and scale morphology, and by sequencing (rDNA), it was possible to see the evolution of scale morphology map on to trees. This marriage of molecular and morphological data made it possible to view distinct groups of species that shared scale detail that might have otherwise been overlooked had either method been used alone. This research has shed significant light on how scale morphology can be used as reliable taxonomic marker for protists, the insights of which can be applied to make taxonomic improvements to other silica-scaled protist groups.

Exercise-induced muscle soreness is characterized by stiffness, tenderness and pain during active movements and weakness of the affected musculature the days after unusually or particularly heavy work. The most pronounced subjective symptoms do not arise immediately but rather between a couple of hours to some days after the exercise (a delayed-onset of muscle soreness), the intensity of pain is greatest about 48 hours after the work. A particular association exists between muscle soreness and eccentric contractions. Despite the fact that muscle soreness is a well known phenomenon in the sphere of sports as well as working life, the pathophysiological mechanisms underlying this are still not understood. In the present study a detailed analysis of human muscle fibre population structure after high tension work (eccentric exercise) that gave rise to muscle soreness, was carried out. The objective was to elucidate how fibres of different types are influenced by repeated muscle contractions reaching extreme tension levels using qualitative and quantitative light and electron microscopic techniques. It was hoped that such morphological analysis would provide a basis for discussion of possible causes for muscle soreness. The muscle function after the work was measured by isokinetic methods. To improve the basis for the ultrastructural analysis the fibre populations in untrained and en­durance trained human m. vastus lateralis of age-matched individuals were classified into different fibre type groups according to their ultrastructure. The selective glycogen depletion from Type 1 fibres seen after long term submaximal work, visualized electron microscopically with PA-TSC-SP staining, substantiated the usefulness of the appearance of the M-band to differentiate between fibre types. Stereological data showed that neither volume density of mitochondria nor of lipid droplets provide sufficient criteria to differentiate between fibre types. After an eccentric exercise regimen sore muscles (m. soleus or m. vastus lateralis) showed disturb­ances of the cross striated band pattern. Fibres with disorganized myofibrillar material made up 1/3, 1/2 and 1/10 of the analysed material, 1 hour, 3 and 6 days after exercise, respectively. The myofibril­lar lesions were preferably localized in the Z-band. This showed streaming, broadening and sometimes total disruption. The Type 2 fibres were most affected. The reduction of strength was greatest with the most rapid contractions. Strength remained de­creased the period when the structural damage was most pronounced. Eight weeks of eccentric muscle training reduced all the above negative effects. The results indicate that the Z-disc constitute the weak link in the myofibrillar contractile chain at high muscle tensions. It is suggested that the myofibrillar lesions are a direct result of mechanical tearing. Rupture of myofibrils is thought to result in formation of protein components and a con- sequental release of protein bound ions that via osmosis result in oedema and soreness. Training, using eccentric contractions over a long period of time leads to adaptations at the fibre level by a reorgani­zation of the contractile apparatus as well as an optimization of nervous coordination. / <p>S. 1-40: sammanfattning, s. 41-79: 5 uppsatser</p> / digitalisering@umu

A deficiência de ferro é a carência nutricional mais frequente no mundo, uma vez que a população de crianças é uma das que mais sofre com essa condição. É sabido que o desenvolvimento cerebral é determinado não apenas por um plano genético, mas sim por uma forte interação de fatores genéticos e ambientais. Evidências emergentes sugerem que a estimulação precoce pode oferecer grande eficácia terapêutica, uma vez que o cérebro é notavelmente responsivo a essa interação com o ambiente. Dado que a estimulação tátil (TS) tem sido previamente demonstrada ser uma abordagem terapêutica eficaz e com potencial aplicação em seres humanos, o objetivo deste estudo foi verificar se a exposição à estimulação tátil desde o dia pós-natal (P) 1 até P32 durante 3 min/dia, poderia ser utilizada para prevenir alterações estruturais do nervo óptico de ratos mantidos com uma dieta deficiente em ferro durante o desenvolvimento pós-natal. Foram utilizados 72 ratos machos recém-nascidos (Wistar), sendo que as ratas-lactantes foram mantidas com dieta isocalórica com 35mg/Fe por kg de ração (Grupo ANTS) ou com 4mg/Fe por kg de ração (Grupo DNTS) durante todo o período de lactação e os filhotes receberam a dieta de suas respectivas ratas-lactantes após o desmame (P22-32). Metade dos filhotes de cada grupo foi submetida à TS diária (Grupo ATS e DTS), durante todo o período experimental (P01-32). Foram realizadas análises estrutural e ultraestrutural, em 3 diferentes idades, para avaliar a integridade tecidual e também a fim de determinar se as mudanças observadas na citoarquitetura do nervo óptico foram significativamente diferentes entre os grupos e idades. Verificou-se que os animais mantidos com dieta deficiente em ferro apresentaram baixo peso corporal a partir do desmame, revelando uma curva de crescimento menos acentuada. A baixa concentração de hemoglobina e hematócritos indicam que esses animais sofreram com anemia severa em todos os períodos estudados. A análise ultraestrutural qualitativa mostrou que a deficiência de ferro imposta durante o período crítico do desenvolvimento leva a sérios danos as fibras das células ganglionares da retina, com efeitos sobre o envoltório de mielina que frequentemente apresentou afrouxamento lamelar e em idades mais avançadas foram encontradas degenerações mielínicas e axonais. A análise ultraestrutural quantitativa mostrou que a dieta deficiente em ferro leva a um atraso no processo de mielinização que pode ser parcialmente revertido pelo tratamento com estimulação tátil. Além disso, fica claro que as fibras de menor diâmetro são mais sensíveis às lesões geradas pela deficiência de ferro e também ao tratamento, enquanto que as fibras de maior diâmetro são afetadas de forma desproporcional tanto pela deficiência de ferro quanto pela estimulação tátil. As lesões sugerem que a transmissão dos sinais elétricos pode estar prejudicada, interferindo com as funções normais do sistema visual. / Iron deficiency has a critical impact on maturational mechanisms of the brain and the damage related to neuroanatomical parameters is not satisfactorily reversed after iron replacement. However, emerging evidence suggest that enriched early experience may offer great therapeutic efficacy in cases of nutritional disorders postnatally, since the brain is remarkably responsive to its interaction with the environment. Given the fact that tactile stimulation (TS) treatment has been previously shown to be an effective therapeutic approach and with potential application to humans, here we ask whether exposure to TS treatment, from postnatal day (P) 1 to P32 for 3 min/day, could also be employed to prevent neuroanatomical changes in the optic nerve of rats maintained on an iron-deficient diet during brain development. It was verified that the animals maintained with iron deficient diet presented low weight from the weaning, revealing a lower growth curve. The low concentration of hemoglobin and hematocrits indicate that these animals suffered from severe anemia in all studied periods. The qualitative ultrastructural analysis showed that the iron deficiency imposed during the critical period of development leads to serious damage to the fibers from retinal ganglion cells, with effects on the myelin sheath that frequently presented lamellar loosening and myelin degenerations and axonal at more advanced age. Quantitative ultrastructural analysis has shown that the iron deficient diet leads to a delay in the myelination process that can be partially reversed by treatment with tactile stimulation. In addition, it is clear that the smaller diameter fibers are more sensitive to the iron deficiency and also to the treatment, whereas the larger fibers are disproportionately affected by both iron deficiency and tactile stimulation. The lesions suggest that the transmission of electrical signals may be impaired, interfering with the normal functions of the visual system.

Através da meniscectomia no lado esquerdo, seguida ou não de enxerto, objetiva-se verificar as possíveis alterações nas superfícies articulares da ATM. Os animais formaram os grupos: GI (meniscectomia); GII (meniscectomia e enxerto); GIII (somente o acesso cirúrgico) que originaram subgrupos conforme o período pós-operatório (10 e 30 dias) e lado (direito e esquerdo). Nos animais GIII, GId, GIId, a Cm e a superfície articular da fossa mandibular (Fm) estavam organizadas em camadas com o predomínio de fibras colágenas do tipo I. Alterações morfológicas como erosão da Cm e Fm e amplos cistos subcondrais no osso subjacente foram verificadas nos animais GIe e GIIe. Em GIIe, a Cm apresentou um nível razoável de organização sob o enxerto. Fibras colágenas do tipo I constituíam as camadas articular e condroblástica nos animais GIe e GIIe e fibras do tipo III predominaram na camada pré-condroblástica. Amplos espaços vasculares eram evidentes na zona bilaminar dos animais GI e GII. / The morphological features of the temporomandibular joint (TMJ) of rats were studied after meniscectomy in the left side of the joint. The specimens were submitted or not to an interpositional auricular cartilage graft. The animals formed the following groups: GI (meniscectomy); GII (meniscectomy and graft); GIII (sham). The post-operative periods (10, 30 days) and the side allowed to form the subgroups. The articular surface of the TMJ exhibited organized layers (GIII, GId, GIId groups) where the type I collagen fibers arranged in regular nets prevailed. Morphological changes as erosion of the articular surface with large subchondral cysts in the subjacent bone were verified (GIe, GIIe). The condyle\'s surface of GII was regular and structurally preserved under the graft. The type I collagen fibers constituted the articular and chondroblastic layers in the GIe and GIIe animals where the type III collagen fibers were predominant in the prechondroblastic layer. Wide vascular spaces were evident in the bilaminar zone of the GI and GII animals.

Este estudo apresenta um painel de dados hematológicos, bioquímicos, morfológicos e citoquímicos para um grupo de serpentes dos gêneros Bothrops e Crotalus mantidas em cativeiro no Instituto Butantan. Para tanto, amostras de sangue foram colhidas da veia coccígea ventral e processadas de acordo com métodos padronizados. Os resultados alcançados foram analisados para determinar diferenças interespecíficas e sexuais. A avaliação citoquímica incluiu Sudan Black B (SBB), Ácido Periódico de Schiff (PAS) e Benzidina Peroxidase (BP). Os resultados hematológicos e bioquímicos mais relevantes indicaram um número significativamente mais alto de basófilos em B. jararaca e níveis mais altos de cálcio nas fêmeas da maioria das espécies. Os azurófilos marcaram positivamente para todas as colorações. A diferenciação entre trombócitos e linfócitos foi facilmente obtida com PAS. Os basófilos marcaram positivamente apenas com PAS em Bothrops alternatus. A marcação em heterófilos variou consideravelmente entre as espécies. Ultra-estruturalmente, os leucócitos foram semelhantes ao já descrito em literatura com algumas variações no tipo dos grânulos dos basófilos e heterófilos. Os grânulos dos basófilos apresentaram-se redondos em sua maioria, com tamanho e densidade variadas, enquanto que os nos heterófilos foram heterogêneos em forma, tamanho e densidade. / This study reports a panel for hematological, biochemical, morphological and cytochemical data for a group of captive snakes belonging to the genus Bothrops and Crotalus mantained at Instituto Butantan, São Paulo, Brazil. Blood samples were collected from ventral coccigeal vein and were processed according to standard protocols. Cytochemical staining including Sudan Black B (SBB), Periodic Acid-Schiff (PAS) and Benzidine peroxidase (BP) were conducted. Blood values were evaluated to determine interspecific and sex differences. We found a significant increase of basophils in Bothrops jararaca and higher levels of calcium in females. Azurophils stained positively for all stains and a differentiation between lymphocytes and thrombocytes was easily obtained with PAS stain. Basophils stained positively only with PAS in Bothrops alternatus. Heterophils staining varied between species. The ultrastructure of leucocytes were similar within described in literature, with some variations on granules of basophils and heterophils. Basophils granules were round, with heterogeneous size and density; whereas, heterphils granules were heterogeneous in shape, size and density.

Avaliou-se a tipificação das fibras musculares e determinou-se parâmetros morfoquantitativos dos efeitos da imobilização por 2 e 4 semanas. Após o período experimental, os músculos (mm.) foram corados por HE e Picro-sírius. Para a tipificação das fibras, utilizou-se as reações da ATPase e NADH-tr; a análise ultraestrutural procedeu-se com microscopia eletrônica de transmissão. Nos resultados, alguns mm. imobilizados (Im) exibiram núcleos centralizados e fibras tipo IIB com formas elusivas. O perimísio destes mm. se espessou, sendo constituído por fibras colágenas tipo III. Os fusos musculares apresentaram espessamento da cápsula. Houve alterações nas linhas Z e modificação de outras ultraestruturas como as mitocôndrias. A área de secção transversa dos mm. Im por 4 semanas foi menor. As fibras do tipo IIB reduziram a área de perfil e aumentaram a densidade, além disso, se ampliou a porcentagem das fibras do tipo IIA. Portanto, a imobilização promoveu alterações nos fusos e esqueleto fibroso muscular, assim como transformou fibras do tipo IIB em IIA. / The effects of muscle immobilization for 2 and 4 weeks were studied by determining morphoquantitative parameters and muscle fiber-typing. After the trial period the muscles (mm.) were subject to HE and Picrosirius stain. The reactions of ATPase and NADH-tr were used for muscles fiber-typing. The ultrastructural analyses were made by transmission electron microscopy. The results showed changes in some of immobilized (Im) mm., it exhibited centralized nuclei cells and type IIB fibers took indefinite forms. The perimisium of Im mm. thicker up, and consisted of type III collagen fibers. Also the muscle spindles capsule thickened. Changes in the Z lines and other ultrastructures such as mitochondria were noticed. The cross-section area of Im mm. for 4 weeks was lower. The type IIB fibers reduced the profile area and density increased. Also the percentage of type IIA fibers expanded. Therefore the immobilization promoted changes in muscle spindles, fibrous skeleton morphology and provide an interconversion of muscle fibers type IIB into IIA.

A gestação inicial da égua é um período fascinante que abrange numerosas e intensas mudanças em seu desenvolvimento, muitas das quais são únicas para a espécie equina. Esse desenvolvimento depende da manutenção da função lútea, do estabelecimento de um ambiente uterino e de uma interação precisa e orquestrada entre o concepto e o ambiente uterino. O objetivo deste estudo foi verificar as alterações histológicas do endométrio e a produção histotrófica em éguas cíclicas e prenhes nos dias 7, 10 e 13 pós-ovulação. No primeiro ciclo, biópsias endometriais de 30 éguas foram coletadas no dia 7 (n = 10), 10 (n = 10) e 13 (n = 10) constituindo o grupo éguas cíclicas. No segundo ciclo, as mesmas éguas foram cobertas por um garanhão fértil, acompanhadas diariamente até detectar a ovulação, considerada o dia 0. Foram coletadas biópsias endometriais nos dias7 (n 10), 10 (n 10) e 13 (n 10). Imediatamente após a coleta, o útero foi lavado e as éguas em que foi obtido embrião, foram inseridas no grupo de éguas prenhes. Um maior calibre dos vasos sanguíneos foi observado em prenhez comparados às éguas cíclicas do dia 7 aos 13. No sétimo dia pós-ovulação, uma grande perda de células ciliadas foi evidente no grupo de éguas prenhes, comparadas ao grupo de éguas cíclicas, as células do epitélio endometrial estavam mais protusas e uma pequena quantidade de secreção histotrófica entre as dobras endometriais foi observada. No décimo dia de prenhez, secreção histotrófica glandular e do epitélio luminal estavam mais presentes comparadas às éguas do grupo cíclico. No dia 13 de prenhes, foi observado um grande conteúdo de histotrofo nas aberturas glandulares que estavam cercadas por células ciliares. Ocorreram alterações no ambiente uterino logo após a entrada do embrião no útero. No estroma e no lúmen, essas modificações parecem visar fornecer a nutrição necessária para o desenvolvimento inicial do embrião e estas mudanças nas estruturas celulares irão interagir na sinalização embrionária, futura fixação, implantação e placentação. / The early pregnancy of mare is a fascinating period that encompasses numerous and intense changes in its development, many of which are unique to the equine species. This development depends on the maintenance of the luteal function, the establishment of a favorable uterine environment and a precise and orchestrated interaction between the concept and the uterine environment. The aim of this study was to evaluate histological changes in the endometrium in days 7, 10 and 13 post-ovulation in pregnant and cyclic mares. In the first cycle, endometrial biopsies from 30 cyclic mares (Cyclic group) were collected on days 7, 10 and 13 post-ovulation. In the second cycle, the same mares were bred by a fertile stallion. At days 7, 10 and 13 post-ovulation intrauterine biopsies were collected. Immediately after sample collection, the mare‟s uteri were flushed, and those mares with embryo recovery were assigned to the Pregnant group. A larger blood vessel caliber was observed in pregnant mares than in cyclic from day 7 to 13. On the 7th day a large loss of ciliated cells was evident in the group of pregnant mares in comparison with the Cyclic group and the superficial cells of the endometrium were more protruded, and a small amount of histotrophic material between the folds was observed. On the 10th day of pregnancy, the glandular histotrophic secretion and the secretion of luminal epithelium became more intense than the secretion of cyclic mares. On the 13th day of pregnancy, a very large amount of histotroph was observed within large glandular openings surrounded by ciliated cells. Changes occurred in the uterine environment thereupon the entry of the embryo into the uterus. In the stroma and in the lumen, these modifications seem aim to provide the necessary nutrition for the initial development of the embryo and to promote changes at cellular structures that will interact in the embryonic signaling and future fixation, implantation and placentation.

Made available in DSpace on 2016-03-22T17:27:02Z (GMT). No. of bitstreams: 1 is2.pdf: 1552188 bytes, checksum: 842b31ea5e16c999c9c802e21d14ccce (MD5) Previous issue date: 2013-07-22 / Tinea nigra is a rare superficial mycosis caused by Hortaea werneckii. This infection presents as asymptomatic brown to black macule mostly in palmo-plantar regions. This study presents dermatoscopy examination of a lesion, which demonstrated a homogeneous nonmelanocytic pigmented pattern with spicules in the macula; scanning electron microscopy (SEM) examination of the fungal culture, showing sympodial conidiogenesis; and SEM examination of a sample of the lesion, that revealed the epidermis with keratinocytes, elimination of fungal filaments and important aggregation of hyphae. SEM s findings correlated with those of dermatoscopic examination and allowed also documenting the mode of dissemination of tinea nigra, showing how hyphae are eliminated on lesion s surface / Tinea nigra é uma rara micose superficial causada pelo fungo Hortaea werneckii. Esta infecção apresenta-se como mancha assintomática acastanhada ou enegrecida, mais frequentemente na região palmo-plantar. Foram realizados neste estudo: dermatoscopia de uma lesão, a qual demonstrou um padrão homogêneo de pigmentação nãomelanocítica com espículas na mácula; microscopia eletrônica de varredura (MEV) da cultura fúngica, mostrando conidiogênese simpodial; e MEV de uma amostra da lesão, a qual revelou epiderme com queratinócitos, eliminação de filamentos fúngicos e importante agregação de hifas. Os achados de MEV se correlacionaram com aqueles do exame dermatoscópico e ainda permitiram documentar o modo de disseminação da tinea nigra, demonstrando como as hifas são eliminadas na superfície da lesão

Apocynaceae presents the flowers with the highest degree of synorganization among the eudicots and highly elaborated pollination mechanisms associated with the high diversity of glands. The osmophore stands out as responsible to produce a floral scent which attracts pollinators and, despite its fundamental relevance for pollination, its structure and mechanism of production and release of the perfume is essentially unknown in Apocynaceae. This present work aims to characterize morphologically and ultrastructurally the osmophores of Apocynaceae, besides chemically identifying the compounds that constitute the floral scent. Species from two subfamilies were selected to describe the diversity of osmophores and types of scent in the family. The osmophores were firstly located histochemically and, later, this region was processed for transmission electron microscopy. Micromorphological analysis was performed by scanning electron microscopy, and the identification of volatile oils made by GC-MS. Osmophores are located on the adaxial surface of the free portion of the petals. They varied in the shape of epidermal cells, striation of the cuticle and presence of trichomes. This gland is mostly formed by secretory epidermis and parenchyma, except in Plumeria, where the osmophores are exclusively epidermal. The secretory cells presented thick walls in the Asclepiadoideae and secretion produced by plastids and SER in all species. Several vesicles perform the intercellular transport of secretion, as well as its release to the environment. However, Plumeria and Ditassa transfer the secretion produced to the vacuole before releasing it, and Tabernaemontana has a mixed release process. The composition of the scent varied among species, with great production of hydrocarbons, alcohols, ketones or monoterpenes depending on the species. Apocynaceae presents a high morphological and metabolic diversity in the osmophores which cannot be correlated with their morphology, subcellular organization, period of secretion release or pollination syndrome. This is the first comprehensive structural study of osmophores in this family that points out to very distinct evolutionary processes that may be related to multiple emergences in the phylogeny and speciesspecific associations with pollinators / Apocynaceae apresenta as flores com o maior grau de sinorganização das eudicotiledôneas e mecanismos de polinização altamente elaborados associados à mais alta diversidade de glândulas. O osmóforo destaca-se como responsável pela produção do perfume floral para atração dos polinizadores e, a despeito de sua importância fundamental para a polinização, a sua estrutura e mecanismo de produção e liberação do perfume é praticamente desconhecido nas Apocynaceae. O presente trabalho tem o propósito de caracterizar morfológica e ultraestruturalmente os osmóforos de Apocynaceae, além de identificar quimicamente os compostos que constituem o perfume floral. Espécies de duas subfamílias foram selecionadas, visando descrever a diversidade de osmóforos e tipos de perfume na família. Primeiramente, os osmóforos foram localizados histoquimicamente e, posteriormente, essa região foi processada para microscopia eletrônica de transmissão. Análise micromorfológica foi realizada através de microscopia eletrônica de transmissão. Análise micromorfológica foi realizada através de microscopia eletrônica de varredura e a identificação dos óleos voláteis, através de GC/MS. Os osmóforos estão localizados na superfície adaxial da porção livre das pétalas. Eles variaram quanto ao formato das células epidérmicas, ornamentação da cutícula e presença de tricomas. Esta glândula é, em sua maioria, formada por epiderme e parênquima secretores, exceto em Plumeria, onde os osmóforos são exclusivamente epidérmicos. As células secretoras apresentaram paredes espessas nas Asclepiadoideae e secreção produzida pelos plastídeos e REL em todas as espécies. Diversas vesículas realizam o transporte intercelular da secreção, assim como a sua liberação para o meio externo. Contudo, Plumeria e Ditassa transferem a secreção produzida para o vacúolo antes de liberá-la e Tabernaemontana possui processo de liberação misto. A composição do perfume variou entre as espécies, havendo grande produção de hidrocarbonetos, alcoóis, cetonas ou monoterpenos dependendo da espécie. Apocynaceae apresenta uma alta diversidade morfológica e metabólica em seus osmóforos que não pode ser correlacionada à morfologia, organização subcelular, período de liberação da secreção ou síndrome de polinização. Esse é o primeiro estudo estrutural abrangente sobre osmóforos na família e aponta para processos evolutivos muito distintos que podem estar relacionados a múltiplos surgimentos na filogenia e associações espécie-específicas com os polinizadores