Effects of processing on structural, mechanical and biological properties of collagen-based substrates for regenerative medicine.

The aim of this work was to investigate the structural features of type I collagen isoforms and collagen-based films at atomic and molecular scales, in order to evaluate whether and to what extent different protocols of slurry synthesis may change the protein structure and the final properties of the developed scaffolds. Wide Angle X-ray Scattering data on raw materials demonstrated the preferential orientation of collagen molecules in equine tendon-derived collagens, while randomly oriented molecules were found in bovine skin collagens, together with a lower crystalline degree, analyzed by the assessment of FWHM (Full Width at Half Maximum), and a certain degree of salt contamination. WAXS and FT-IR (Fourier Transform Infrared) analyses on bovine collagen-based films, showed that mechanical homogenization of slurry in acidic solution was the treatment ensuring a high content of super-organization of collagen into triple helices and a high crystalline domain into the material. In vitro tests on rat Schwannoma cells showed that Schwann cell differentiation into myelinating cells was dependent on the specific collagen film being used, and was found to be stimulated in case of homogenization-treated samples. Finally DHT/EDC crosslinking treatment was shown to affect mechanical stiffness of films depending on collagen source and processing conditions.

A simple approach to synthetize folic acid decorated magnetite@SiO2 nanostructures for hyperthermia applications.

Superparamagnetic magnetite nanoparticles were synthetized and capped by a SiO2 shell in order to avoid oxidation and aggregation of the iron oxide nanostructures. The inorganic capping was then further decorated by folic acid molecules, by using a very simple procedure exploiting supramolecular interactions among the organic moieties and the inorganic nanoparticles. The supramolecular nanoadduct thanks to folic acid molecules could act as a "Trojan horse" for the cancer cells and due to its superparamagnetic properties could induce local heat generation upon an appropriate magnetic field application. In fact, temperature was increased up to 42 °C when a 18 mT magnetic field was applied to the nanoparticles and the hybrid nanostructures were verified to be selectively internalized by HeLa cells, a human cervical cancer line known to overexpress the folic acid receptor.

Cell proliferation and growth-associated protein 43 expression in the olfactory epithelium in Poecilia reticulata after copper solution exposure.

This study investigated the regeneration in the olfactory mucosa of the teleostean fish Poecilia reticulata when returned to dechlorinated tap water after 4-day exposure to 30 microg/L of Cu(2+). The regeneration process in the olfactory tissue was examined in fishes at 0, 3, 6 and 10 days of recovery in well water. Jade B staining permitted to evaluate the rate of the damage which was especially extended to olfactory neurons. Immediately after the end of exposure, a massive mitotic activity in the basal region of the mucosa was detected by immunostaining with PCNA. After 3 days of recovery the nuclei of the newly formed cells had already finished their migration to the upper portion of the epithelium, and cellular division was much less intense. Simultaneously, immunoreactivity for the neural growth-associated phosphoprotein GAP-43 increased respect to control levels, revealing that the new differentiating PCNA-positive elements belonged to immature neurons. After 6 days in well water no mitotic activity was detected, while the GAP-43 labelling appeared particularly concentrated in the apical surface of the olfactory epithelium. After 10 days the aspect of the olfactory epithelium was almost identical to the control. The present results suggest that after 10 days regeneration seems to be complete and integrity of the tissue restored. Furthermore, the epithelium reconstitution does not show apparent divergence from other fishes or mammals.

Leishmaniasis in Tuscany (Italy): (I) An investigation on phlebotomine sandflies in Grosseto Province.

A survey of phlebotomines has been made in the area of the Province of Grosseto (Tuscany, Italy) where human cases of both visceral and cutaneous leishmaniasis have recently occurred. 12 collecting stations, placed around the leishmaniasis foci, were divided into three groups of localities, the Argentario Promontory, coastal plains territory and hilly inland territory. Sandflies were collected with lit and unlit oil traps, and by hand. A total of 11,032 specimens (57% males) were caught and identified. Of these, 86.6% were Phlebotomus perfiliewi, 10.8% P. perniciosus and 2.3% Sergentomyia minuta; only a few specimens were P. papatasi and P. mascittii. In the first group of localities, P. perniciosus was very abundant (87.8%) and S. minuta scarce (12.0%); in the second group, P. perfiliewi was the only species collected; in the third group, P. perfiliewi was very abundant (98.0%), S. minuta was vert scarce (1.1%) and P. perniciosus even more so (0.7%). P. perniciosus is thought to be the probable vector of visceral leishmaniasis in the area of the Monte Argentario and P. perfiliewi is strongly suspected of playing a role in transmission of the cutaneous form in the third group of localities. The species responsible for transmitting visceral leishmaniasis in some of the foci in this area is yet to be found.

Leishmaniasis in Tuscany (Italy): (II) Leishmania form wild Bodentia and Carnivora in a human and canine leishmaniasis focus.

From April 1977 to February 1978, wild mammals were trapped in a focus of low leishmanial endemicity (Baccinello) in the Province of Grosseto (Tuscany, Italy). Spleen homogenates of the trapped animals were injected i.p. into hamsters, from which spleen biopsies were taken after 60 days and impression smears carefully observed for the presence of Leishmania. Positive spleens were subinoculated into hamsters. 404 animals of 12 species were trapped: 31 insectivores, 317 rodents and 56 carnivores. Black rats (Rattus rattus) and field mice (Apodemus sylvaticus) were the rodent species most frequently trapped--160 and 139 respectively--of which 143 and 134 respectively were tested in hamsters for Leishmania. 35 foxes (Vulpes vulpes) were caught, of which 16 were tested for Leishmania. Four hamsters were found infested with Leishmania. Three of them had been injected with spleen homogenates from R. rattus and one from V. vulpes although liver and spleen impression smears of all trapped animals were negative for Leishmania on direct microscopical examination. The significance of Leishmania infection in wild animals in the leishmanial focus of Baccinello is discussed.

Studies on Mediterranean leishmaniasis: V.A. preliminary epidemiological survey of human leishmaniasis in Tuscany.

Leishmanin skin testing was performed on 1,285 people in the Tuscany region of Italy. In a known endemic area for leishmaniasis there was an increasing positivity with age, while in another area the curve was bell-shaped suggesting an interruption of transmission in the recent past. In all areas the positivity rate for males was higher than for females, but it was not possible to distinguish between areas of cutaneous and visceral disease.

Leishmaniasis in Tuscany (Italy): (IV) An analysis of all recorded human cases.

Recent investigations on the distribution of human and canine leishmaniasis, and on the vectors and wild reservoirs of the disease in foci in Tuscany have stimulated a retrospective evaluation of human leishmaniasis of the whole region. From a study of the literature, of statistical reports (ISTAT) and of unpublished information, 77 cases of visceral leishmaniasis and 97 of cutaneous leishmaniasis have been analysed. From the case reports of both forms it appears that: (a) a higher incidence of cases occurred in restricted areas; (b) visceral and cutaneous leishmaniasis have been hypoendemic in Tuscany since the years of the earlier reports (1933 and 1936, respectively); (c) the number of infantile visceral cases was much higher in the period from 1933 to 1955 (84.5% of subjects in the 0-10 years age group) than in the subsequent years, 1956-75 (50.0%). The possible significance of the infantile and adult forms of visceral leishmaniasis in Tuscany is discussed, and the local data are compared with those of other regions.

Leishmaniasis in Sardinia: II. Natural infection of Phlebotomus perniciosus Newstead, 1911, by Leishmania infantum Nicolle, 1908, in the province of Cagliari.

Two promastigote-infected specimens of Phlebotomus perniciosus Newstead, 1911, were found in the province of Cagliari, Italy. The parasites from one specimen were successfully cultured in a hamster. The isolate was typed on 11 enzymes and shown to be identical with the reference strain of Leishmania infantum s.st. This is the first typing of a leishmanial organism from naturally infected P. perniciosus.

Leishmaniasis in Tuscany (Italy): VII. Studies on the role of the black rat, Rattus rattus, in the epidemiology of visceral leishmaniasis.

The recent finding that four Leishmania isolates from Rattus rattus are enzymologically identical to L. infantum, aetiological agent of human and canine visceral leishmaniasis in the Mediterranean area, has stimulated further research on the role of R. rattus in the epidemiology of visceral leishmaniasis in Tuscany, Italy. The present work has led to the following results: (a) R. rattus experimentally infected with L. infantum displayed a general resistance to the parasite, even though it appeared that a very low number of amastigotes persisted in the animal for a long period; (b) Phlebotomus perniciosus and P. perfiliewi induced to feed on rats, with either a massive visceral infection or a subpatent immunodepressed infection, became infected, though in different proportions; (c) P. perniciosus and P. perfiliewi were readily attracted to, and fed on, the black rat in nature. These findings, together with those obtained earlier, strongly suggest that the black rat may be one of the wild reservoirs of L. infantum in the areas under study.

Leishmaniasis in Sardinia. 5. Leishmanin reaction in the human population of a focus of low endemicity of canine leishmaniasis.

About half (640 subjects) of the population of Soleminis, a canine leishmaniasis focus 15 km north of Cagliari, Sardinia, was subjected to the leishmanin skin test. The total positive rate was 9.7%, and that of males was 15.5%. The age trend in the positive rate resembled a hyperbolic curve, with the 0-5 years age group rate being zero and the highest rate (39.3%) occurring in the 55-65 years age group. The Soleminis focus was classified as hypoendemic, with a prevalence of canine leishmaniasis of about 3%. Dog surveillance, land reclamation and changes in population habits during the last few decades are considered to be responsible for the reduction of transmission to man.

Leishmaniasis in Tuscany (Italy): VI. Canine leishmaniasis in the focus of Monte Argentario (Grosseto).

The results of a survey of canine leishmaniasis in the coastal focus of Monte Argentario (Tuscany, Italy) are reported. Of the 171 dogs examined, 41 (23.9%) were positive in the fluorescent antibody test (FAT), but only 12 (7.0%) were positive by microscopic observation of lymphatic ganglia. Of the 41 FAT positive dogs, 17 (10.0%) showed signs of the disease. The evolution of leishmaniasis over one year from the first examination showed that 88.0% of the dogs with signs died of the disease, and 36.0% of the asymptomatic dogs became patent or died; 52.0% of the asymptomatic dogs had apparently recovered. The prevalence of the disease, the geographical, age, sex and race distribution of cases, the evolution of leishmaniasis in dogs and FAT positivity are discussed.

[Zooprophylaxis: old and new problems]. / Zooprofilassi: un problema vecchio e attuale.

In the present note the authors report on the most important works which have examined the application of zooprophylaxis as a means for malaria control from the onset of this century to the present day. The method of zooprophylaxis has evolved throughout the years in close connection with other problems, undoubtedly more important in the history of malaria, which in the present paper are amply discussed. From the very beginning this subject was the object of heated debates by those who retained it a valid method of malaria control and those who denied its validity, among whom some of the most important Italian and foreign malariologists. The experiments of zooprophylaxis received most attention in the twenties, but, following the works of Missiroli and co-workers in the early thirties by which the varieties of the "complex maculipennis" were identified, the interest on zooprophylaxis was less heightened. It should be noted, however, that the increase of cattle, and especially land reclamation, reduced considerably malaria endemicity in northern Italy where zoophilic varieties were most prevalent. Recently, renewed interest on zooprophylaxis has been shown in Pakistan, where some experiments on the use of cattle have led to an unexpected increase of malaria transmission.

Philippo Bonanni and his "Micrographia curiosa".

In this introduction, a short biography of Philippo Bonanni and comments on his "Micrographia curiosa" are presented. P. Bonanni was the first naturalist to describe a phlebotomine sandfly. His work was published in 1691, and on the occasion of the Tricentenary of its publication, the "First International Symposium on Phlebotomine Sandflies" was organized.

Studies on mating plug of two sandfly species, Phlebotomus perniciosus and Phlebotomus papatasi (Diptera: Psychodidae).

A study was undertaken to interpret the nature and function of the "plug like formation" observed inside the spermathecae of many phlebotomine sandflies dissected during field surveys. Trials were carried out on two laboratory reared species, Phlebotomus perniciosus and P. papatasi. The results showed that the "plug like formation" is a true mating plug (MP) containing immotile sperms. Our studies showed also that the MP evolves differently according to the physiological state of the mated female. In blood-fed females, it persists 22-25 hrs post mating. Follows a period of 3-4 days in which the spermathecae acquire different form from that of the virgin spermathecae, but sperms are still undistinguishable. Active sperms appear only a few hours before oviposition. After oviposition, however, spermathecae are void as in the virgin female. Thus, during the complete gonotrophic cycle the spermathecae display five different forms: a) normal and void; b) showing MP; c) distended and yellowish, MP absent; d) active sperms present; e) normal and void. In unfed females, MP was observed to last for 6 days post copula. The results show also that MP is interfering with reinsemination.

Drastic nickel ion removal from aqueous solution by curcumin-capped Ag nanoparticles.

A completely green synthesis protocol has been adopted to obtain silver nanoaggregates capped by the natural compound (1E, 6E)-1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-diene), also known as curcumin. The synthesis has been monitored by infrared, Raman, visible and fluorescence spectroscopies. Characterization confirms that curcumin reduces and caps the nanoparticles, and such a procedure allows its solubility in water and drastically increases curcumin stability. Silver nanoparticles (AgNPs)/curcumin complex has been dispersed in a water solution containing a known nickel ion concentration. After three days, a grey precipitate is observed and nickel concentration in the solution is reduced by about 70%.