An in vivo humanized model to study homing and sequestration of Plasmodium falciparum transmission stages in the bone marrow

IntroductionRecent evidence suggests that the bone marrow (BM) plays a key role in the diffusion of P. falciparum malaria by providing a “niche” for the maturation of the parasite gametocytes, responsible for human-to-mosquito transmission. Suitable humanized in vivo models to study the mechanisms of the interplay between the parasite and the human BM components are still missing.MethodsWe report a novel experimental system based on the infusion of immature P. falciparum gametocytes into immunocompromised mice carrying chimeric ectopic ossicles whose stromal and bone compartments derive from human osteoprogenitor cells.ResultsWe demonstrate that immature gametocytes home within minutes to the ossicles and reach the extravascular regions, where they are retained in contact with different human BM stromal cell types.DiscussionOur model represents a powerful tool to study BM function and the interplay essential for parasite transmission in P. falciparum malaria and can be extended to study other infections in which the human BM plays a role

Exploring mobility in Italian Neolithic and Copper Age communities

As a means for investigating human mobility during late the Neolithic to the Copper Age in central and southern Italy, this study presents a novel dataset of enamel oxygen and carbon isotope values (δ18Oca and δ13Cca) from the carbonate fraction of biogenic apatite for one hundred and twenty-six individual teeth coming from two Neolithic and eight Copper Age communities. The measured δ18Oca values suggest a significant role of local sources in the water inputs to the body water, whereas δ13Cca values indicate food resources, principally based on C3 plants. Both δ13Cca and δ18Oca ranges vary substantially when samples are broken down into local populations. Statistically defined thresholds, accounting for intra-site variability, allow the identification of only a few outliers in the eight Copper Age communities, suggesting that sedentary lifestyle rather than extensive mobility characterized the investigated populations. This seems to be also typical of the two studied Neolithic communities. Overall, this research shows that the investigated periods in peninsular Italy differed in mobility pattern from the following Bronze Age communities from more northern areas

Le fasi neolitica ed eneolitica di Moscano di Fabriano (AN)

Recent excavations at Moscano in Fabriano have uncovered a double inhumation grave in a reused cavity, for which two 14C dates have been identified. The grave was found on the outskirts of a Copper Age settlement, featuring rough surface pottery similar to that discovered on layer 4 at Attiggio in Fabriano. The two burials at Moscano lay one above the other, with a thin layer of soil separating them. The lower inhumation is an incomplete female adult skeleton (Individual 1) and was probably laid in a huddled position. A flint arrowhead was probably part of the grave goods. The upper burial is of a young individual aged around 8 years (Individual 1A) laid out in a supine position. It is not clear whether a pottery fragment decorated with scales and found at the site is related to this individual. This burial is currently unique for several reasons, including its location in inland Marche, the laid out position of the upper body and the presence of pottery decorated with scales. Although problematic in several respects, the 14C dates of the two individuals are suggestive of end of 4th / early 3rd millennium BC. This would make these discoveries contemporary to the most recent burials at Fontenoce and the central phase of the settlement at Conelle in Arcevia.Recenti scavi hanno riportato alla luce a Moscano di Fabriano una duplice sepoltura per la quale si è in possesso di datazioni. La sepoltura, rinvenuta ai margini di un abitato eneolitico con ceramica a superficie scabra molto simile a quella dello str. 4 di Attiggio di Fabriano, era all’interno di una cavità riutilizzata a scopo funerario. Le due deposizioni sono sovrapposte, separate da un sottile strato di terreno. L’inferiore è uno scheletro incompleto di un individuo femminile di età adulta (individuo 1) deposto probabilmente in posizione semirannicchiata al quale è da riferire una punta di freccia. La superiore, completa, è di un individuo subadulto di circa 8 anni (individuo 1A) inumato in posizione supina, la cui relazione con un frammento ceramico a squame non è del tutto certa. La tomba al momento rappresenta un unicum per vari elementi (il rinvenimento nell’area interna delle Marche, la posizione distesa dello scheletro superiore e la probabile relazione con la ceramica rusticata). Le datazioni dei due individui, pur presentando alcune problematiche, si collocano tra la fine del IV millennio e l’inizio del III millennio a.C., quindi contemporanei alle sepolture più recenti di Fontenoce e alla fase centrale di vita dell’insediamento di Conelle di Arcevia

THE ANATOMY OF BONE SIALOPROTEIN IMMUNOREACTIVE SITES IN BONE AS REVEALED BY COMBINED ULTRASTRUCTURAL HISTOCHEMISTRY AND IMMUNOHISTOCHEMISTRY

Bone sialoprotein was immunolocalized at the EM level in thin Lowicryl K4M sections of rat bone. Because of the unconventional EM morphology of the bone matrix seen in thin demineralized acrylate sections, the pattern of immunolabeling was compared with detailed structural images of demineralized bone obtained using an en bloc treatment of tissue samples with the cationic electron 'dye,' Malachite Green (MG), which provides stabilization and retention of anionic material throughout specimen processing. A system of structures corresponding to the sites of bone sialoprotein (BSP) immunoreactivity, as seen in Lowicryl K4M thin sections, could be readily identified in the MG-treated, epoxy thin sections. This system includes the cement lines, and aggregates of similar material within mineralized bone and mineralizing osteoid. The virtual identity of BSP distribution with the arrangement of the MG-visualized material indicates that a BSP-enriched, noncollagenous phase can be demonstrated using different, unrelated tissue preparation and imaging protocols for EM. Besides improving our understanding of the distribution of bone sialoprotein in bone, these data assign a previously unrecognized structural dimension to noncollagenous material in the bone matrix

Localization of bone sialoprotein (BSP) to Golgi and post-Golgi secretory structures in osteoblasts and to discrete sites in early bone matrix.

Bone sialoprotein (BSP), a bone matrix-enriched glycoprotein containing the Arg-Gly-Asp (RGD) motif and endowed with cell binding properties, was localized in osteoblasts and early bone matrix of developing rat bone at the ultrastructural level. Preliminary light microscopic observations indicated that intracellular labelling was restricted to a paranuclear dot corresponding to the "negative Golgi image" of classical histology. The same pattern was observed whether antisera against the fully glycosylated protein or a peptide antiserum to a stretch of amino acids in human BSP sequence were employed. At the EM level, we obtained labeling over the Golgi area of osteoblasts but not over the rER. The labeling was concentrated over distensions of the trans Golgi and over pro-secretory granules. In the matrix, BSP was distributed in a non-random manner. The label was concentrated over spherical aggregates of finely fibrillar material corresponding to the sites of early mineral deposition (so-called "mineralization nodules"). Such BSP-positive foci were seen both close to and away from the cell surface. The predominant association of BSP with Golgi and post-Golgi secretory structures and its absence from rER, as well as the reproducibility of the same pattern of localization with different antisera, might indicate a slow transit of the protein through the Golgi, not necessarily associated with protein glycosylation

Dissection of the human bone marrow environment as a privileged niche for Plasmodium falciparum gametocyte development

Plasmodium falciparum gametocytes develop in the human host in 10-12 days and only mature stage V can be found in the peripheral circulation. Observations from histological studies of a systematic organ survey in pediatric cases of fatal malaria (1) and from an analysis of bone marrow samples from anemic children infected by P. falciparum (2) revealed that immature gametocytes accumulate in the human bone marrow and that they are readily observed in the extravascular sites of this organ, altogether putting the human bone marrow under the spotlight as a privileged niche for gametocyte maturation and for having a key role in human-to-mosquito transmission of the malaria parasites. The mechanism(s) driving gametocyte sequestration in the human bone marrow and which parasite sexual stage is involved in homing are unclear. As in vitro systems recapitulating the complexity of human bone marrow are presently missing, a Bone Marrow Humanized Mouse (BMHM) model based on osteoprogenitor cell transplantation has been established (3) and recently refined (4) to reproduce a microenvironment for marrow structural development and for suitable for hematopoiesis. P. falciparum transgenic lines producing fluorescent gametocytes, have been used in the BMHM model to investigate gametocyte-BM interactions obtaining preliminary results on parasite (i) vascular or extravascular distribution, (ii) sequestration timing, (iii) stage(s) involved