Unraveling the role of mechanical stimulation on smooth muscle cells: A comparative study between 2D and 3D models.

A thorough understanding of cell response to combined culture configuration and mechanical cues is of paramount importance in vascular tissue engineering applications. Herein, we investigated and compared the response of vascular smooth muscle cells (vSMCs) cultured in different culture environments (2D cell monolayers and 3D cellularized collagen-based gels) in combination with mechanical stimulation (7% uniaxial cyclic strain, 1 Hz) for 2 and 5 days. When cyclic strain was applied, two different responses, in terms of cell orientation and expression of contractile-phenotype proteins, were observed in 2D and 3D models. Specifically, in 2D configuration, cyclic strain caused ∼50% of cell population to align nearly perpendicular (80-90 degrees) to the strain direction, while not influencing the contractile-phenotype protein expression, as compared to the 2D static controls. Conversely, the application of uniaxial strain to 3D constructs induced a ∼60% cell alignment almost parallel (0-10 degrees) to the strain direction. Moreover, 3D mechanical stimulation applied for 5 days induced a twofold increase of SM α-actin level and a 14-fold increase of calponin expression as compared to 3D static controls. Altogether these findings provide a new insight into the potential to drive cell behavior by modulating the extracellular matrix and the biomechanical environment. Biotechnol. Bioeng. 2016;113: 2254-2263. © 2016 Wiley Periodicals, Inc.

Chondrocyte response to high regimens of cyclic hydrostatic pressure in 3-dimensional engineered constructs.

PURPOSE: Despite widespread use of 3-dimensional (3D) micro-porous scaffolds to promote their potential application in cartilage tissue engineering, only a few studies have examined the response to hydrostatic pressure of engineered constructs. A high cyclic pressurization, currently believed to be the predominant mechanical signal perceived by cells in articular cartilage, was used here to stimulate bovine articular chondrocytes cultured in a synthetic 3D porous scaffold (DegraPol). METHODS: Construct cultivation lasted 3 days with applied pressurization cycles of amplitude 10 MPa, frequency 0.33 Hz, and stimulation sessions of 4 hours/day. RESULTS: At 3 days of culture, with respect to pre-culture conditions, the viability of the pressurized constructs did not vary, whereas it underwent a 16% drop in the unpressurized controls. Synthesis of alfa-actin was 34% lower in all cultured constructs. Synthesis of collagen II/collagen I did not vary in pressurized constructs, was 76% lower in unpressurized controls, and was around 230% higher in pressurized constructs with respect to unpressurized controls. Chondrocytes showed a phenotypic spherical morphology at time zero and at 3 days of pressurized culture. CONCLUSIONS: Although the passage from 2D expansion to 3D geometry was effective to guide cell differentiation, only mechanical conditioning enabled the maintenance and further cell differentiation toward a mature chondrocytic phenotype.

Probing lysine mono-methylation in histone H3 tail peptides with an abiotic receptor coupled to a non-plasmonic resonator.

Binder and effector molecules that allow studying and manipulating epigenetic processes are of biological relevance and pose severe technical challenges. We report the first example of a synthetic receptor able to recognize mono-methylated lysines in a histone H3 tail peptide, which has relevant functions in epigenetic regulation. Recognition is robust and specific regardless of the position and the number of mono-methylated lysines along the polypeptide chain. The peptide is first captured in solution by a tetraphosphonate cavitand (Tiiii) that selectively binds its Lys-NMe+ moieties. Separation from solution and detection of the peptide-Tiiii complexes is then enabled in one single step by an all dielectric SiO2-TiO2 core-shell resonator (T-rex), which captures the complex and operates fully reproducible signal transduction by non-plasmonic surface enhanced Raman scattering (SERS) without degrading the complex. The realized abiotic probe is able to distinguish multiple mono-methylated peptides from the single mono-methylated ones.

Gene therapy: The state of the art and future directions.

Gene therapy, because of its aim to eradicate the causes rather than the symptoms of diseases, is believed by many to be the therapy of the future. The problems of developing clinically viable gene therapeutic approaches and designing safe and efficient gene delivery reagents are inseparable: shortcomings in one are going to affect adversely the success of the other. It is generally accepted that the major impediment to the successful application of gene therapy for the treatment of a range of dis-eases is not a paucity of therapeutic genes, but the lack of efficient non-toxic gene delivery systems. Transfection vectors com-monly used in gene therapy are mainly of two types: viral and non-viral. Non-viral gene delivery is currently the subject of increasing attention because of its relative safety and its simplicity of use; however, its use is still far from being ideal due to its comparatively low efficiency. The purpose of this review is to give an overview of the current knowledge concerning the assembly of lipoplexes and the trafficking of lipoplexes into cells, as well as to underscore the advantages and disadvantages of lipidic gene carriers among non-viral gene delivery systems.

Data in support of Gallium (Ga(3+)) antibacterial activities to counteract E. coli and S. epidermidis biofilm formation onto pro-osteointegrative titanium surfaces.

This paper contains original data supporting the antibacterial activities of Gallium (Ga(3+))-doped pro-osteointegrative titanium alloys, obtained via Anodic Spark Deposition (ASD), as described in "The effect of silver or gallium doped titanium against the multidrug resistant Acinetobacter baumannii" (Cochis et al. 2016) [1]. In this article we included an indirect cytocompatibility evaluation towards Saos2 human osteoblasts and extended the microbial evaluation of the Ga(3+) enriched titanium surfaces against the biofilm former Escherichia coli and Staphylococcus epidermidis strains. Cell viability was assayed by the Alamar Blue test, while bacterial viability was evaluated by the metabolic colorimetric 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay. Finally biofilm morphology was analyzed by Scanning Electron Microscopy (SEM). Data regarding Ga(3+) activity were compared to Silver.

Tumor necrosis factor alpha and glutathione interplay in chronic heart failure.

The pro-inflammatory cytokine, tumor necrosis factor alpha (TNF alpha), in concert with neurohormones, contributes to chronic heart failure (CHF) progression. This implies that TNF a antagonism may constitute an important target for CHF therapy. However, clinical trials in CHF patients using compounds that trap TNF alpha, comprising infliximab, an antibody directed to TNF alpha, and etanercept, a soluble recombinant receptor of TNF alpha, gave disappointing results bringing back to light the dual, short-term beneficial and long-term harmful effect of TNF alpha. This review focuses on the dual, concentration- and time-related effects of TNF alpha, the yin and yang action of TNF alpha in cardiac ischemia/reperfusion and contraction. Importantly, the harmful effects of TNF a are related to glutathione deficiency, a common hallmark to several other chronic inflammatory diseases. Recently, in rat models of CHF, oral administration of the glutathione precursor, N-acetylcysteine (NAC), was shown to hinder pathways of TNF alpha harmful signalling and to rescue cardiac structure and function. These results suggest that glutathione deficiency in association with TNF alpha activation may play a role in the pathophysiology of CHF and that NAC may represent a potential therapy in CHF.

Synthesis and antibacterial activity of a series of basic amides of teicoplanin and deglucoteicoplanin with polyamines.

Basic carboxamides of teicoplanin A2 (CTA) and its aglycon (TD) are prepared by condensation of the 63-carboxyl function of these antibiotics with linear or branched polyamines. The antimicrobial activities of some of the resulting compounds were better than those of the unmodified antibiotics. The presence of more than one basic group in the amidic chain enhanced the in vitro activity of some TD-amides against Gram-negative bacteria; two of these derivatives were also effective in vivo against Escherichia coli septicemia in the mouse. Among the CTA derivatives, the amide with spermine showed some unexpected in vitro activity against Gram-negatives. Both CTA- and TD-amides with polyamines are very soluble in water over a wide range of pH and are very hydrophilic.

The morphology of early trophoblast after dioxin poisoning in the Seveso area.

The placental morphology in 22 cases of induced abortion performed one to three months after the Seveso disaster is described. In 15 cases no gross histological or ultrastructural alterations were observed. In seven cases there were many prominent microprecipitates beneath the trophoblastic basement membrane at various distances from the fetal capillaries. Intermediate patterns between simple dense bodies, already well known, and complex pluri-stratified giant structures not previously reported have been described. Combined ultrastructural and x-ray investigations (EDAX analysis) have recorded calcium- and iron-related emissions from these structures. The significance of these findings and their possible relationship to dioxin poisoning has been discussed.

Double uterus, blind hemivagina, and ipsilateral renal agenesis: 36 cases and long-term follow-up.

OBJECTIVE: To review the experience of the Milan University First Department of Obstetrics and Gynecology in patients with double, didelphic, of bicornuate uterus, blind hemivagina, and ipsilateral renal agenesis, and to consider the frequently unsatisfactory surgical approach. METHODS: Thirty-six women with double, didelphic, or bicornuate uterus, blind hemivagina, and ipsilateral renal agenesis were identified from clinical records for the period 1962 to 1992. We evaluated demographic data, disease, symptoms, correctness of therapeutic approach, and definitive treatment. RESULTS: Seventeen patients previously had undergone incomplete surgery in other hospitals and 19 were treated by us for the first time. Total hysterectomy was performed on two of the 36 women and hemihysterectomy and hemicolpectomy were performed on four. In the other 30, the vaginal septum was excised and marsupialization was done. The pregnancy rate in the 15 women wanting children was 87% and the live birth rate was 77%. Serial biopsy specimens were obtained from the lateral fornix after the excision of the septum in 13 of the non-hysterectomized patients over 1-9 years and revealed progressively more extensive areas of squamous metaplasia of müllerian epithelium. In some isolated cases, papillary hyperplasia, mild dysplasia, and vaginal adenosis were found. At the end of follow-up, 16 patients still did not want children. Follow-up was possible in 34 cases. CONCLUSION: Early accurate diagnosis after menarche followed by excision and marsupialization of the blind hemivagina offers complete relief of symptoms and preserves reproductive potential. Partial morphologic changes are evident but metabolic modifications comparable to those of the adjacent normal vagina have not yet been documented.

Ectopic pregnancy and recurrent spontaneous abortion: two associated reproductive failures.

The association between spontaneous abortion and ectopic pregnancy was evaluated in a case-control study conducted on 161 women (cases) with recurrent spontaneous abortions (two or more consecutive spontaneous abortions) and 170 control subjects who delivered normal infants. The risk of ectopic pregnancy in women with a history of recurrent spontaneous abortion was about fourfold that of controls (relative risk adjusted for age and number of pregnancies = 4.3; 95% confidence interval 1.4-14.7). This association was confirmed by comparing the observed number of extrauterine pregnancies in women with recurrent spontaneous abortions with the expected number computed from regional data on the frequency of ectopic pregnancies; the estimated relative risk was 3.7, with a 95% confidence interval of 2.2-7.0. The present report found an association between spontaneous abortions and ectopic pregnancies, suggesting some common risk/etiologic factors for these two reproductive failures.

Serum CA 125 measurements in the diagnosis of endometriosis recurrence.

Seventy-one women underwent follow-up laparoscopy for persistent infertility six to 24 months after conservative treatment of endometriosis. The disease was staged according to the revised American Fertility Society classification of 1985. The serum CA 125 concentration was also measured in each patient to evaluate its efficacy in the diagnosis of endometriosis recurrence. The patients with stages I and II endometriosis had serum CA 125 levels not significantly higher than in the patients with negative findings, whereas those with stages III and IV endometriosis presented significantly higher levels (P less than .005 and P less than .001, respectively) than the disease-free women. The sensitivity of serum CA 125 measurements in the diagnosis of endometriosis recurrence was 14.8%, the specificity was 100%, and the predictive values of normal (less than 35 U/mL) and elevated levels were 27 and 100%, respectively. In some cases, serum CA 125 measurements may be used instead of follow-up laparoscopy, or to indicate when laparoscopy should be performed or postponed.

Ultrasound in the diagnosis of subclasses of unicornuate uterus.

Fourteen infertile women with hysterographic diagnoses of unicornuate uterus underwent pelvic ultrasonography and, subsequently, laparoscopy to evaluate the ability of ultrasound to diagnose the subclasses of this malformation. Seven women underwent laparotomy to remove a rudimentary horn. Ultrasound demonstrated a sensitivity of 85.7% and a specificity of 100% in diagnosing the presence of a rudimentary horn, and a sensitivity of 80% and a specificity of 100% in diagnosing the presence of a cavity in the rudimentary horn. Urography confirmed the supplementary information on the urinary tract provided by ultrasound. Ultrasound appears to have a role in the clinical management of women with unicornuate uterus.

Magnetic resonance evaluation of double uteri.

A total of 18 infertile patients with hysterosalpingographic diagnosis of bifid uterus underwent magnetic resonance imaging (MRI) and subsequent laparoscopy or laparotomy to evaluate the capability of MRI to differentiate among the various classes of the malformation. Magnetic resonance imaging identified both bicornuate uteri correctly, both didelphic uteri, nine of the 12 partial septate uteri, and both complete septate uteri. The method demonstrated a sensitivity of 100% and a specificity of 78.6% in demonstrating cervical prolongation of the spur. Compared with laparoscopy, MRI is less expensive and less invasive, and can also be performed in patients with extensive adhesions. However, it has the disadvantage of not providing information on tubal conditions or on the presence of minimal and mild endometriosis.

Endometrial reconstruction after hysteroscopic incisional metroplasty.

Endometrial reconstruction after hysteroscopic metroplasty of a septate uterus was studied by light microscopy and scanning electron microscopy when the uterus was removed 13 days after metroplasty for a histologic diagnosis of leiomyosarcoma. We observed a retraction of the margins of the incised septum, which could prevent the formation of intrauterine adhesions. Reepithelialization of the cut surface seemed to proceed both centripetally, by proliferation of the surrounding normal endometrium, and centrifugally from the bottom of the glands present at the base of the septum.

Repetitive conservative surgery for recurrence of endometriosis.

We evaluated the recovery of fertility and the relief of pain symptoms in a long-term follow-up of 42 women undergoing repetitive conservative surgery for recurrent endometriosis. The mean age of the patients was 31.1 +/- 4.3 years. At the time of their second operation the disease was stage IV in 14 women, stage III in 25, and stage I in three. After reoperation, the patients were followed for a mean period of 41.8 +/- 30.3 months. Pain symptoms returned in eight women, dysmenorrhea and deep dyspareunia in eight, and pelvic pain in seven. Eight of the 28 women (28.6%) who attempted to conceive achieved a total of 13 pregnancies. The corrected pregnancy rate was 35%, and the cumulative rate at 27 months was 30.7%. A third operation was necessary in six women after a mean period of 35 months. Conservative surgery is an effective therapeutic option for infertile patients with recurrent endometriosis.

Inguinal endometriosis: pathogenetic and clinical implications.

In six women with a preoperative diagnosis of incarcerated hernia, surgical exploration of the groin revealed inguinal endometriosis and no hernia. The affected structure was always the extraperitoneal portion of the round ligament. Six of the seven lesions were on the right. Intraperitoneal endometriosis was demonstrated in all patients. Catamenial pain was the pathognomonic symptom in the differential diagnosis of the inguinal mass. Gynecologists and surgeons should bear in mind the possibility that endometriosis may be the cause of symptoms of a suspected incarcerated inguinal hernia. Should this disease be detected at inguinal exploration, a laparoscopy is indicated during the same operation.

Magnetic resonance imaging in Mayer-Rokitansky-Küster-Hauser syndrome.

Six girls with suspected Rokitansky syndrome underwent magnetic resonance imaging (MRI) and then sonography and laparoscopy to confirm the clinical diagnosis and define the precise characteristics of the malformation. The uterus was missing in all; four had müllerian rudiments in place of the uterus, which were cavitary in two as shown by sonography and in one by MRI. Both ovaries were present in all cases, normally located in four and extrapelvic in two; they were correctly identified by MRI in all cases and by sonography in four. Partially canalized vaginal rudiments were recognized by MRI in two girls and by sonography in one. One patient had unilateral renal agenesis, demonstrated by both MRI and sonography. Magnetic resonance imaging was more precise than laparoscopy and sonography in defining the anatomical characteristics of Rokitansky syndrome, and is less invasive and expensive than laparoscopy.

Endometrium of cavitary rudimentary horns in unicornuate uteri.

The rudimentary horns removed from ten patients with unicornuate uterus, subclass A1b, were studied by light and scanning electron microscopy. Seven of the horns removed were small in volume (range 7.5-15.7 mL) and three were larger (range 20.8-39.7 mL). In four cases the endometrial cavity communicated with the tubal lumen, hematometra was present in three cases, and endometriosis was diagnosed in six. In six of the seven rudimentary horns without hematometra, the endometrial mucosa did not correspond to the cycle phase of the endometrium of the hemiuterus; however, in the three rudimentary horns with hematometra, the endometrium presented cycle-phase characteristics similar to those of the hemiuterus. The endometrio-myometrial junction, the underlying myometrium, and the arcuate arterioles were always altered. We consider that the absence of cyclic changes in the endometria of the smaller horns may be attributed to reduced vascular supply, to the absence of any isthmocervical structure, and to the qualitative and/or quantitative differences in endometrial mucosal receptors.

Endometriosis and nonobstructive müllerian anomalies.

We reviewed the case records of all women (N = 748) who underwent laparoscopy for infertility in our department from 1985-1990. The patients were divided into two groups, cases and controls, according to the presence of nonobstructive müllerian anomalies (N = 198) or absence of such anomalies (N = 545); women with obstructive müllerian anomalies (N = 5) were excluded. The patients were analyzed for the presence of endometriosis, malformation class, and productive history. The frequency of endometriosis was 30.8% in the women with nonobstructive müllerian anomalies, versus 38.5% in the controls (P = .209). Among the subjects with unicornuate uterus, the prevalence of endometriosis (55%) was significantly greater compared with the patients with other nonobstructive müllerian anomalies (28%) (P less than .05), but not compared with the controls (38.5%). Overall, no differences were observed in the frequency of endometriosis between infertile women with and without nonobstructive müllerian anomalies. Our results seem to exclude a common pathogenetic factor underlying endometriosis and nonobstructive müllerian anomalies.

Laparoscopic ovarian puncture for correct staging of endometriosis.

Small, deep ovarian endometriomas are not easily diagnosed. In 52 infertile patients, laparoscopy demonstrated enlarged ovaries (maximum diameter 3.5 to 5 cm) with a smooth whitish surface and no mature follicles, corpora lutea, or other cysts. Ovarian puncture was performed, and endometriotic material aspirated in 25 women (48.0%). Cyst diameter was calculated using the geometric formula r = 3 square root of 3V/4 pi where r = radius and V = volume of liquid aspirated. Eight patients with apparently normal pelvis had endometriosis, and 14 with apparent minimal or mild endometriotic lesions were restaged. Laparoscopic ovarian puncture of enlarged ovaries was important for correct diagnosis and staging of endometriosis.