The impact of SARS-CoV-2 pandemic on Oncologic and Degenerative Spine Surgery Department activity: the experience of Rizzoli Orthopaedic Institute under COVID-19 lockdown.

OBJECTIVE: Experience of Department of Oncologic and Degenerative Spine Surgery of Rizzoli Orthopaedic Institute during SARS-CoV-2 pandemic lockdown. PATIENTS AND METHODS: Retrospective observational study of surgically treated patients from 09th March 2020 to 04th May 2020. DATA COLLECTED: age, sex, type of disease, neurological status, days of hospitalization, complications and type of discharge. A comparison analysis with same period of the last year was performed in order to evaluate the impact of COVID-19 spreading on daily surgical activity. RESULTS: A total of 107 surgical procedures in 102 patients were performed from 09th March 2020 to 04th May 2020. Analysis showed a statistically significant difference in age, sex, ASIA class and type of treated disease compared to the same period of the last year (p=0.042, 0.006, 0.022 and 0.007, respectively). No statistically significant differences were observed in type of discharge, length of hospitalization and complications (p= 0.447, 0.261 and 0.127, respectively). 3 COVID-19 infections have been identified in hospitalized patients. 1 COVID-19 patient wad admitted from Emergency Department and was managed according to a dedicated path. CONCLUSIONS: Surgical activity was paradoxically increased during SARS-CoV-2 pandemic lockdown through the management of urgent and non-deferrable spinal disease with a low rate (3,9%) of COVID-19 infections.

Percolation thresholds in ultrasound compacted tablets.

Twenty matrix systems with different KCl content (as drug model, from 10 to 90% w/w) and Eudragit RS-PM (as inert excipient) were prepared using an ultrasound-assisted press and a traditional eccentric machine. The release behavior from both types of matrices was examined; the kinetic parameters for the release (intrinsic dissolution) and the technological properties of the final tablets (total porosity) were used to estimate the percolation threshold for the drug model and the excipient in both systems. For the systems compacted by ultrasound (US) the estimated value for the excipient percolation threshold ranges from 13.4 to 20.2% v/v (lower than that found for traditional tablets), that agrees with a continuum percolation model suggesting the presence of a continuum phase inside the tablet. This depends on a thermoplastic deformation of Eudragit RS-PM under ultrasound, that destroyed the particulate system of the excipient and transform it into a continuum medium. The percolation threshold for KCl ranged from 58.6 to 61.0% v/v for US and from 26.7 to 42.2% v/v for the traditional tablets. The higher value for ultrasound compacted tablets can be explained by the difficulty of KCl to outcome from a matrix containing insoluble phase that surrounds KCl crystals.

Fractal analysis of beta-cyclodextrin-indomethacin particles compacted by ultrasound.

An association between indomethacin and beta-cyclodextrin (beta-CD) was obtained by compacting a 1:2 molar physical mixture by ultrasound. The product prepared by this technique was compared with the initial physical mixture and with materials having the same composition but prepared by a simple compaction and kneading process. The samples examined by scanning electron microscopy revealed morphological differences related to the methods of preparation. In particular the material obtained by ultrasound had a smooth surface and on milling produced particles of uniform size; moreover EDAX analysis (energy dispersion analysis by X-rays) also revealed a homogeneous distribution of the two components in each particle examined. Surface fractal dimension of the surface of these particles is very low, suggesting the presence of a regular and smooth surface whereas the sample obtained by kneading had a higher value. The reactive dimension related to dissolution was much higher than the corresponding surface dimension in all cases and had comparable values for all samples regardless of the techniques used. This behavior was attributed to the hydrophilicity of beta-CD, which levels the differences during dissolution and which could be originated by surface imperfections. Experimental results suggested that the material obtained by ultrasound had a dissolution rate comparable to that measured with the kneaded material. The use of ultrasound, however, can both reduce the production time and improve the homogeneity of the association between indomethacin and beta-CD.

Effects of ultrasound-assisted compaction on Ketoprofen/Eudragit S100 mixtures.

Ketoprofen alone and in binary mixtures with Eudragit S100 was compacted by an ultrasound-assisted (US) tableting machine at an energy ranging from 50 to 400 J. The final material was analysed by TLC and HPLC: no decomposition product of the active agent was found. IR spectra and HSM revealed the absence of any interaction between the two components. Thermal analysis (DSC) evidenced that ketoprofen inside the mixtures was transformed into an amorphous state, documented by the decreasing of the DeltaHfus as the Eudragit/ketoprofen ratio increases and as US energy increases. While pure ketoprofen recovers its crystalline state quickly after the US treatment, the presence of Eudragit was found to slow down or possibly to prevent the regeneration of the crystallinity.

Ultrasound-compacted and spray-congealed indomethacin/polyethyleneglycol systems.

The product obtained by ultrasound (US)-assisted compaction was compared with a solid dispersion for systems containing polyethyleneglycols (PEGs) of different molecular weights and indomethacin (IMC), at the weight ratio 9:1, obtained by traditional melting and followed by a new US-assisted spray-congealing technique. US-discharge during compaction affects crystallinity of both IMC and PEG: pure IMC changes to an amorphous form and, when in mixture with PEG, partially dissolves in the excipient: this causes an increase of the dissolution rate of the drug. Differential scanning calorimetry (DSC) thermograms do not reveal any endothermic peak associated with the melting of the drug, while X-ray diffractograms show a loss of crystallinity of both IMC and PEG in the US-compacted granules. The extent of a back-crystallisation, which reduces the dissolution rate, as a function of the ageing of the material, depends on the type of the selected PEG. When a molten IMC/PEG mixture was transformed into microspheres by an US-assisted spray-congealing technique, the behaviour at dissolution almost recalls that of US-compacted granulates and some differences are briefly discussed.

Description and preliminary evaluation of a new ultrasonic atomizer for spray-congealing processes.

A new atomizer that operates with ultrasonic energy is described. This apparatus is intended to obtain microparticulate drug delivery systems through spray-congealing or spray-drying technologies. In this work, some experimental results are reported on model systems submitted to spray-congealing. The formulations under examination contained theophylline and fenbufen as model drugs and stearic acid, carnauba wax, Cutina HR(R) and Compritol 888 ATO(R) as low melting excipients. Non-aggregate and spherical-shaped microparticles were obtained with all the materials tested; moreover, they had smooth surface and good flowability. The particle sizes depend on the amount of drug present and in each case the maximum size value of the distribution frequency was found to be 375 mu. In vitro release of the drug depends on its solubility and on the excipient lipophilicity. The results suggest that the ultrasound-assisted atomizer could be proposed as a possible alternative to traditional atomizers used for spray-congealing in the pharmaceutical field.

Formation of ion-pairs in aqueous solutions of diclofenac salts.

In this work we studied the ability of the diclofenac anion to form ion-pairs in aqueous solution in the presence of organic and inorganic cations: ion-pairs have a polarity and hydrophobicity more suitable to the partition than each ion considered separately and can be extracted by a lipid phase. The cations considered were those of the organic bases diethylamine, diethanolamine, pyrrolidine, N-(2-hydroxyethyl) pyrrolidine and N-(2-hydroxyethyl) piperidine; the inorganic cations studied were Li(+), Na(+), K(+), Rb(+), Cs(+). Related to each cation we determined the equilibrium constant (K(XD)) for the ion-pair formation with the diclofenac anion in aqueous solution and the water/n-octanol partition coefficient (P(XD)) for each type of ion-pair formed. Among the alkali metal cations, only Li(+) shows some interaction with the diclofenac anion, in agreement with its physiological behaviour of increasing clearance during the administration of diclofenac. The influence of the ionic radius and desolvation enthalpy of the alkali metal cations on the ion-pair formation and partition was briefly discussed. Organic cations promote the formation of ion-pairs with the diclofenac anion better than the inorganic ones, and improve the partition of the ion-pair according to their hydrophobicity. The values of the equilibrium parameters for the formation and partition of ion-pairs are not high enough to allow the direct detection of their presence in the aqueous solution. Their formation can be appreciated in the presence of a lipid phase that continuously extracts the ion-pair. Extraction constants (E(XD)=P(XD) times K(XD)) increase passing from inorga to organic cations. This study could help to clarify the mechanism of the percutaneous absorption of diclofenac in the form of a salt, a route where the formation of ion-pairs appears to play an important role.

Preparation and characterization by morphological analysis of diclofenac/PEG 4000 granules obtained using three different techniques.

The steam granulation is a new wet granulation technique, which involves the use of steam water instead of traditional liquid water as granulation liquid. The aim of this work was to evaluate the possibility of using this new technique to prepare diclofenac-polyethylene glycol 4000 accelerated-release granules. Steam granules were prepared in a laboratory scale high-shear mixer, and their properties were then compared to those of granules, having the same composition, obtained by traditional granulation techniques (wet and melt granulation). The results showed that, selecting the proper process parameters, it was possible to obtain granules using all the three methods; however, the total process time was significantly shorter for steam granulation (30 min) in comparison to traditional wet granulation (70 min), due to the lower amount of used water. The morphological characterization of steam, water and melt granules, performed by scanning electron microscopy (SEM) and image analysis, revealed that steam granules had a more spherical shape and a larger surface area with respect to water and melt ones, suggesting a possible difference in dissolution behavior. Moreover, differential scanning calorimetry (DSC) and X-ray powder diffraction analysis evidenced the transformation of the drug from its originally crystalline form into the amorphous one. Finally, the in vitro dissolution tests showed an increased dissolution rate of the drug from the granules (in particular steam granules) in comparison to pure drug and physical mixture. In conclusion, the results of this study suggested that the steam granulation technique could be considered an interesting alternative to traditional wet granulation to improve the dissolution rate of diclofenac.

Role of Lefty in the anti tumor activity of human adult liver stem cells.

Recent studies demonstrated that factors derived from embryonic stem cells inhibit the tumorigenicity of a variety of cancer cell lines. Embryonic stem cell-secreted Lefty, an inhibitor of Nodal-signalling pathway, was implicated in reprogramming cancer cells. Whether adult stem cells exhibited similar properties has not been explored. The aim of the present study was to investigate whether the conditioned medium (CM) derived from adult stem cells influence in vitro and in vivo tumor growth by a Nodal-dependent pathway. In particular we compared the anti-tumor effect of CM from human liver stem cells (HLSC) with that of bone marrow-derived mesenchymal stem cells (MSC). We found that HLSC-CM inhibited the in vitro growth and promoted apoptosis in HepG2 cells that expressed a deregulated Nodal pathway. The effect of HLSC-CM was related to the presence of Lefty A in the CM of HLSC. Silencing Lefty A in HLSC or Lefty A blockade with a blocking peptide abrogated the anti-proliferative and pro-apoptotic effect of HLSC-CM. Moreover, the administration of human recombinant Lefty A protein mimicked the effect of HLSC-CM indicating that Nodal pathway is critical for the growth of HepG2. At variance of HLSC, bone marrow-derived MSC did not express and release Lefty A and the MSC-CM did not exhibited an anti-tumor activity in vitro, but rather stimulated proliferation of HepG2. In addition, the intra-tumor administration of HLSC-CM was able to inhibit the in vivo growth of HepG2 hepatoma cells implanted subcutaneously in SCID mice. At variance, HLSC-CM derived from Lefty A silenced HLSC was unable to inhibit tumor growth. In conclusion, the results of present study suggest that Lefty A may account for the tumor suppressive activity of HLSC as a result of an inhibition of the Nodal-signalling pathway by a mechanism similar to that described for embryonic stem cells.

A novel system for large-scale gene expression analysis: bacterial colonies array.

In the present work, we report the use of bacterial colonies to optimize macroarray technique. The devised system is significantly cheaper than other methods available to detect large-scale differential gene expression. Recombinant Escherichia coli clones containing plasmid-encoded copies of 4,608 individual expressed sequence tag (ESTs) were robotically spotted onto nylon membranes that were incubated for 6 and 12 h to allow the bacteria to grow and, consequently, amplify the cloned ESTs. The membranes were then hybridized with a beta-lactamase gene specific probe from the recombinant plasmid and, subsequently, phosphorimaged to quantify the microbial cells. Variance analysis demonstrated that the spot hybridization signal intensity was similar for 3,954 ESTs (85.8%) after 6 h of bacterial growth. Membranes spotted with bacteria colonies grown for 12 h had 4,017 ESTs (87.2%) with comparable signal intensity but the signal to noise ratio was fivefold higher. Taken together, the results of this study indicate that it is possible to investigate large-scale gene expression using macroarrays based on bacterial colonies grown for 6 h onto membranes.

Availability of NSAIDH beta-cyclodextrin inclusion complexes.

The diffusion of a series of non steroidal antiinflammatory drugs through a silicone rubber membrane has been studied from suspensions both of the free and beta-cyclodextrin complexes forms at different pH values of the medium. Higher diffusion rates of the complexed forms as compared with the free ones and a rate-limiting effect of the apparent stability constant in the diffusion of the complexes were observed.

MMH cells: An in vitro model for the study of retinol-binding protein secretion regulated by retinol.

The untransformed stable cell line Met murine hepatocytes (MMH), generated from liver explants of transgenic mice expressing a constitutively active truncated form of the human hepatocyte growth factor receptor (cyto-Met), represents an innovative tool for in vitro studies of liver function. In the present report, we show that the MMH-D3 line isolated from the liver of a 3-day-old mouse is a useful model to investigate the regulation of the synthesis and secretion of retinol-binding protein (RBP) by retinol (vitamin A alcohol). Experiments with Northern blot hybridization, metabolic labeling of cellular proteins followed by immunoprecipitation, and Western blot analysis demonstrated that, similarly to the in vivo situation, in MMH-D3 cells the presence of retinol does not affect transcriptional and translational rate of the RBP gene but is essential for regulating the secretion rate of the protein. Unlike HepG2 human hepatocarcinoma cells used thus far in studies of retinoid metabolism, including the synthesis and secretion of RBP, vitamin A deficiency causes, in MMH-D3 cells, the inhibiton of RBP secretion and the protein accumulation in the cell, whereas retinol repletion promptly results in RBP secretion. This model will be very useful in future studies on vitamin A distribution in the organism.

[The ovary at puberty]. / L'ovaire pubertaire.

Our investigation includes two groups of subjects in which the relationships between ovarian growth, steroidogenesis and gonadotropin secretion were studied. The first group consists of premenarchal girls studied in a cross-sectional program; the second group includes girls after menarche with irregular cycles who were investigated with a long term follow up. In the first group an increase in ovarian volume and in the number of developing follicles were seen with increasing chronological age and pubertal stages. With advancing pubertal development, a progressive increase of LH pulse frequency and amplitude can be observed. These variables appear strongly linked to the ovarian morphological and functional development as the significant correlation between LH mean levels and pulsatile characteristics on the one hand, and ovarian volume and steroid production on the other hand demonstrate. In the maturational stage just before menarche, the ovaries may show increased volume and a very dishomogeneous structure; ovarian steroidogenesis may be often mainly directed towards the androgen synthesis. After menarche ovarian volume and structure continue to change and normal or enlarged ovaries, with homogeneous, multifollicular or polycystic structure can be frequently observed. However normal and homogeneous ovaries prevail in adolescents with regular and ovulatory cycles, while enlarged and dishomogeneous ovaries prevail in irregular anovulatory cycles. A trend towards normal adult ovarian morphology and structure is present in ovulatory subjects with normal gonadotropin pulsatile release and steroid synthesis. On the other hand, deranged gonadotropin profiles imply irregular cycles anovolation, hyperandrogenemia and enlarged polycystic ovaries during adolescence.(ABSTRACT TRUNCATED AT 250 WORDS)