Seeing is believing? When scanning electron microscopy (SEM) meets clinical dentistry: The replica technique.

In restorative dentistry, the in situ replication of intra‐oral situations, is based on a non‐invasive and non‐destructive scanning electron microscopy (SEM) evaluation method. The technique is suitable for investigation restorative materials and dental hard‐ and soft‐tissues, and its interfaces. Surface characteristics, integrity of interfaces (margins), or fracture analysis (chipping, cracks, etc.) with reliable resolution and under high magnification (from ×50 to ×5,000). Overall the current study aims to share detailed and reproducible information about the replica technique. Specific goals are: (a) to describe detailed each step involved in producing a replica of an intra‐oral situation, (b) to validate an integrated workflow based on a rational sequence from visual examination, to macrophotography and SEM analysis using the replica technique; (c) to present three clinical cases documented using the technique. A compilation of three clinical situations/cases were analyzed here by means the replica technique showing a wide range of possibilities that can be reached and explored with the described technique. This guidance document will contribute to a more accurate use of the replica technique and help researchers and clinicians to understand and identify issues related to restorative procedures under high magnification.     

Note: lack of influence of adherent Lactobacillus isolates on the attachment of Escherichia coli to the intestinal epithelial cells of chicken in vitro

Two Lactobacillus isolates, Lact. acidophilus I 26 and Lact. fermentum I 25, were selected, based on their poor aggregation with Escherichia coli and strong ability to adhere to ileal epithelial cells (IEC), to study in vitro interactions with E. coli O1:K1, O2:K1 and O78:K80 in an IEC radioactive-assay under the conditions of exclusion (lactobacilli and IEC, followed by the addition of E. coli), competition (lactobacilli, IEC and E. coli together) and displacement (E. coli and IEC, followed by the addition of lactobacilli). The results indicated that Lact. acidophilus I 26 and Lact. fermentum I 25 could not significantly reduce the attachment of E. coli O1:K1, O2:K1 and O78:K80 to IEC under the three conditions tested in vitro, except that the attachment of E. coli O1:K1 was slightly reduced by Lact. fermentum I 25 in the test for competition.     

Cytological studies on the developing vitreous as related to the hyaloid vessel system

The embryonic development of the cell population of the mammalian vitreous has been traced to two sources: the undifferentiated mesenchymal cells of the eye primordium and the primitive reticular cells of the bone marrow. Undifferentiated mesenchymal cells invade the future vitreous space in two ways: through the annular opening between the rim of the optic cup and the lens primordium, and through the open embryonic fissure. They differentiate into prevascular cells, hemangioblasts, and fibrocytes located in the area of the optic nerve head. From the very beginning of fetal development, another ameboid-type cell of mesenchymal origin makes its entrance into the vitreous through the hyaloid vessels; these monocyte-like cells differentiate into hyalocytes and populate a well-defined area of the cortical vitreous close to the retina and to the ora serrata. Gamma-irradiation (600 rads) of newly born rabbits and cats decreases the number of migrating amebocytes in their vitreous; 24 h later, however, they are replaced by monocytes from the hyaloid vessels.     

A simple breathing circuit minimizing changes in alveolar ventilation during hyperpnoea

Many clinical and research situations require maintenance of isocapnia, which occurs when alveolar ventilation (V'A) is matched to CO2 production. A simple, passive circuit that minimizes changes in V'A during hyperpnoea was devised. It is comprised of a manifold, with two gas inlets, attached to the intake port of a nonrebreathing circuit or ventilator. The first inlet receives a flow of fresh gas (CO2=0%) equal to the subject's minute ventilation (V'E). During hyperpnoea, the balance of V'E is drawn (inlet 2) from a reservoir containing gas, the carbon dioxide tension (PCO2) approximates that of mixed venous blood and therefore contributes minimally to V'A. Nine normal subjects breathed through the circuit for 4 min at 15-31 times resting levels. End-tidal PCO2 (Pet,CO2) at rest, 0, 1.5 and 3.0 min were (mean+/-SE) 5.1+/-0.1 kPa (38.1+/-1.1 mmHg), 4.9+/-0.1 kPa (36.4+/-1.1 mmHg), 5.0+/-0.2 kPa (37.8+/-1.6 mmHg) and 5.0+/-0.2 kPa (37.6+/-1.4 mmHg) (p=0.53, analysis of variance (ANOVA)), respectively; without the circuit, Pet,CO2 would be expected to have decreased by at least 2.7 kPa (20 mmHg). Six anaesthetized, intubated dogs were first ventilated at control levels and then hyperventilated by stepwise increases in either respiratory frequency (fR) from 10 to 24 min(-1) or tidal volume (VT) from 400 to 1,200 mL. Increases in fR did not significantly affect arterial CO2 tension (Pa,CO2) (p=0.28, ANOVA). Only the highest VT decreased Pa,CO2 from control (-0.5 +/- 0.3 kPa (-3.4 +/- 2.3 mmHg), p<0.05). In conclusion, this circuit effectively minimizes changes in alveolar ventilation and therefore arterial carbon dioxide tension during hyperpnoea.     

Interaction of rabbit C-reactive protein with phospholipid monolayers studied by microfluorescence film balance with an externally applied electric field

C-reactive protein (CRP) is one of the most characteristic acute-phase proteins in humans and many other animals. It binds to phosphorylcholine in a calcium-dependent manner. In addition, CRP activates the complement systems via the classical pathway. The interaction between rabbit CRP (rCRP) and model biological membrane is studied using dimyristoylphosphatidylethanolamine and dipalmitoylphosphatidylcholine monolayers. Observations with fluorescence microscopy indicate that rCRP is more likely to be incorporated in the liquid phase of monolayers. Such incorporation does not depend on the presence of calcium and is not inhibited by phosphocholine. The area occupied by the protein when incorporated into the monolayer was estimated. The dipole moment density of the protein crossing the air/water interface was measured by applying an external electric field. Our results indicate that calcium binding leads to a conformational change in CPR, which might modify the orientation of CRP in the monolayer. In addition, a negative charge or negative difference in dipole moment density facilitates the incorporation of CPR into the monolayer.     

Cardiovascular effects produced by microinjection of angiotensins and angiotensin antagonists into the ventrolateral medulla of freely moving rats

In this study we determined the cardiovascular effects produced by microinjection of angiotensin peptides [Angiotensin-(1-7) and Angiotensin II] and angiotensin antagonists (losartan, L-158,809, CGP 42112A. Sar1-Thr8-Ang II, A-779) into the rostral ventrolateral medulla of freely moving rats. Microinjection of angiotensins (12.5-50 pmol) produced pressor responses associated to variable changes in heart rate, usually tachycardia. Unexpectedly, microinjection of both AT1 and AT2 ligands produced pressor effects at doses that did not change blood pressure in anesthetized rats. Conversely, microinjection of Sar1-Thr8-Ang II and the selective Ang-(1-7) antagonist, A-779, produced a small but significant decrease in MAP an HR. These findings suggest that angiotensins can influence the tonic activity of vasomotor neurons at the RVLM. As previously observed in anesthetized rats, our results further suggest a role for endogenous Ang-(1-7) at the RVLM. The pressor activity of the ligands for AT1 and AT2 angiotensin receptor subtypes at the RVLM, remains to be clarified.