Amniotic fluid prostaglandin E2 in pregnancies complicated by preterm prelabor rupture of the membranes
Tomáš Bestvina, MUDr.
bestvinat@lfhk.cuni.cz
Ivana Musilova1, Ctirad Andrys2, Marcela Drahosova2, Helena Hornychova3, Bo Jacobsson4, Ramkumar Menon5, Piotr Laudanski6, Martin Stepan1, Tomas Bestvina1, and Marian Kacerovsky1, 7
1Department of Obstetrics and Gynecology, Charles University in Prague, Faculty of Medicine Hradec Kralove, Czech Republic. 2Department of Clinical Immunology and
Allergy, Charles University in Prague, Faculty of Medicine Hradec Kralove, University Hospital Hradec Kralove, Czech Republic. 3Fingerland´s Department of
Pathology, Charles University in Prague, Faculty of Medicine Hradec Kralove, University Hospital Hradec Kralove, Czech Republic. 4Department of Obstetrics and Gynecology, Sahlgrenska University Hospital, Gothenburg, Sweden. 5Department of
Obstetrics and Gynecology, Division of Maternal-Fetal Medicine & Perinatal Research, The University of Texas Medical Branch at Galveston, Galveston, USA.
6Department of Perinatology, Medical University of Bialystok, Bialystok, Poland
7Biomedical Research Center, University Hospital Hradec Kralove, Czech Republic.
Objective: To determine amniotic fluid prostaglandin E2 levels in women preterm prelabor rupture of the membranes with respect to microbial invasion of the amniotic cavity (MIAC), intraamniotic inflammation (IAI), microbial-associated IAI, histological chorioamnionitis, and short-term neonatal outcome. [1-10]
Methods: One hundred forty-five women with singleton pregnancies were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis and were assayed for prostaglandin E2 levels by ELISA. IAI was defined as amniotic fluid interleukin-6 > 745 pg/mL. Microbial-associated IAI was defined as the presence of both MIAC and IAI.
Result: No differences in prostaglandin E2 levels were found between women with and without MIAC (p=0.27). Women with IAI (with IAI: median 214.8 pg/mL vs.
without IAI: median 113.2 pg/mL; p=0.0008) and microbial-associated IAI (with microbial-associated IAI: median 214.3 pg/mL vs. without microbial-associated IAI:
median 114.5 pg/mL; p=0.01) had higher levels of prostaglandin E2 level than women without these conditions in crude analysis, as well as after adjustment for gestational age at sampling (p<0.0001 for both). Women with histological chorioamnionitis had higher prostaglandin E2 levels only in crude analysis (p=0.02), but not after adjustment for gestational age at sampling (p=0.10). No associations between amniotic fluid prostaglandin E2 concentrations and short-term neonatal morbidity was found.
Conclusions: In conclusion, the presence of IAI or microbial-associated IAI is associated with higher amniotic fluid PGE2 concentrations in women with preterm prelabor rupture of membranes.
References
[1] Gibb W: The role of prostaglandins in human parturition. Ann Med 1998;30(3):
235-241.
[2] Lorenz RP, Botti JJ, Chez RA, Bennett N: Variations of biologic activity of low- dose prostaglandin E2 on cervical ripening. Obstet Gynecol 1984;64(1): 123-127.
[3] Lee SE, Romero R, Park IS, Seong HS, Park CW, Yoon BH: Amniotic fluid prostaglandin concentrations increase before the onset of spontaneous labor at term.
The journal of maternal-fetal & neonatal medicine : the official journal of the European Association of Perinatal Medicine, the Federation of Asia and Oceania Perinatal Societies, the International Society of Perinatal Obstet 2008;21(2): 89-94.
[4] Romero R, Baumann P, Gomez R, Salafia C, Rittenhouse L, Barberio D, et al.:
The relationship between spontaneous rupture of membranes, labor, and microbial invasion of the amniotic cavity and amniotic fluid concentrations of prostaglandins and thromboxane B2 in term pregnancy. Am J Obstet Gynecol 1993;168(6 Pt 1):
1654-1664; discussion 1664-1658.
[5] Menon R, Fortunato SJ, Milne GL, Brou L, Carnevale C, Sanchez SC, et al.:
Amniotic fluid eicosanoids in preterm and term births: effects of risk factors for spontaneous preterm labor. Obstet Gynecol 2011;118(1): 121-134.
[6] Romero R, Emamian M, Wan M, Quintero R, Hobbins JC, Mitchell MD:
Prostaglandin concentrations in amniotic fluid of women with intra-amniotic infection and preterm labor. Am J Obstet Gynecol 1987;157(6): 1461-1467.
[7] Romero R, Wu YK, Mazor M, Hobbins JC, Mitchell MD: Amniotic fluid prostaglandin E2 in preterm labor. Prostaglandins Leukot Essent Fatty Acids 1988;34(3): 141-145.
[8] Leaver HA, MacPherson HD, Hutchon DJ: Amniotic fluid prostaglandins F2 alpha and E2, measured at artificial rupture of the membranes, predict the subsequent progress of labour. Prostaglandins Leukot Med 1987;28(3): 237-242.
[9] Mazor M, Wiznitzer A, Maymon E, Leiberman JR, Cohen A: Changes in amniotic fluid concentrations of prostaglandins E2 and F2 alpha in women with preterm labor.
Isr J Med Sci 1990;26(8): 425-428.
[10] Hsu CD, Meaddough E, Aversa K, Hong SF, Lee IS, Bahodo-Singh RO, et al.:
Dual roles of amniotic fluid nitric oxide and prostaglandin E2 in preterm labor with intra-amniotic infection. Am J Perinatol 1998;15(12): 683-687.
1
Segmental microcirculation in a site of various types of bowel anastomosis in experiment Slavomír Blažej, MD
sblazej@azet.sk
Academic Dpt. of Surgery, Medical Faculty of Charles University, Hradec Králové Co-Authors: J. Páral, Z. Turek, A. Ryška, M. Pavlík, V. Radochová, B. Jegorov, M. Kaška Tutor: Assoc. Prof. Dr. Milan Kaška, MD, PhD
Introduction
Contemporary abdominal surgery is facing an increasing number of malignant diseases of the large bowel. Anatomical conditions together with one’s skill seem to be the limits of operation techniques. Alternatives of digestive tract reconstructions of laden bowel are sought after. Incidence of a malignant disease concerns the large bowel at most. Its surgical therapy is based on resection and reconstruction using common methods such as suture and stapling.
A potentially usable method seems to be the application of a special glue such as one of the non-suture anastomoses that already have more than hundred years’ tradition. The first successful use of metal rings called “buttons” in digestive organs reconnection was carried out by the American surgeon Murphy in the 19th century. But a long time before, similar techniques using available materials such as hollow parts of wood, egg shells, goose-quill, etc.
were applied. Biodegradable synthetic materials entered the market with scientific progress with some complications. So an alternative use of glued bowel reconnection is being examined. Cyanoacrylate has been recently proven to be suitable for that purpose. Now another question has been raised: how significantly glues alter the morphological and physiological status of the glued tissue and if there is a difference between the glued and sutured or stapled tissue blood microcirculation and pato-morphologic condition, respectively.
Microcirculation level is one of the criteria in determining the impact of different techniques of bowel reconnection as blood microcirculation is the limiting factor of anastomotic healing.
Aims and hypothesis
The aim of the experimental study is to confirm whether there is a significant difference in the microcirculation level next to a stapled, sutured or glued large bowel anastomotic site in the pig. Experimental hypothesis claims that glued anastomosis leads to the same load factor of the connected bowel ends tissue as the stapling or suture methods.
Materials and methods
The pilot study included 20 female pigs from the controlled breed of average weight 40 kgs.
Pigs were stabled in hutches of FVZ UO vivarium in the area of FRN Hradec Králové a week before the experiment took place and fed with unitized dry diet and tap water. 24 hrs before the experiment animals were fed only with tap water. On the day of the experiment, the vascular input, premedication, and general anesthesia were performed by a cooperating veterinary doctor. The supine position for tracheal intubation and ventilation was carried out.
Afterwards a cannula was placed into the femoral artery for continuous blood pressure measurement. Afterwards heart frequency and oxygenation of peripheral blood were observed. Flushing out the rectum and standard operation site preparation were next steps.
Lower middle laparotomy and controlled drainage of the urine bladder proceeded. The sigmoid colon was identified and isolated from the rest of organs. Two microcirculation detection probes (Laser Doppler Flowmetry - LDF) were placed close to the anastomotic site and fixed with gentle sewing material (Dafilon 5/0) to serosa. Actual functional capillary density can be evaluated in cm/cm2. Both oral and aboral end probes were placed 20mm from the intended transection and following anastomotic line. The microcirculation status was
2
measured. Scissors were used to transect the bowel following the flush of the aboral intestine end with saline solution and the microcirculation condition was evaluated. Subsequently anastomosis was performed using an absorbable knitted thread for standard sewing technique or a stapler (MSM, 25) or glue (Glubran2 – cyanoacrylic glue, GEM s.r.I., Italy) utilizing an adapted circular stapler from the same manufacturer. Afterwards, microcirculation intensity was measured 60 and 120 min after the construction of anastomosis. Five measurements in 2 second intervals (for the calculation of the mean volume) were performed in the above- mentioned moments according to the experimental time axis. Peritoneal cavity check regarding bleeding or other pathology was performed. Detection probes were removed and laparotomy closed. Skin suture was treated with Akutol. After 6 to 7 days in a hutch with standardized diet and tap water intake, relaparotomy in general anesthesia was performed. A 10cm long biopsy of the large bowel including anastomosis was withdrawn. Animals were spent by exsanguination into the peritoneal cavity in general anesthesia. The biopsy was examined by the pathologist (HE-staining) microscopically focusing on the status of healing anastomosis. Measured values of blood microcirculation status were statistically processed using the software SigmaPlot 13.0.
Results
Of the original number of 20 pigs, 8 animals were disqualified for technical problems during the operation and other significant problems in experimental technique introduction. 12 pigs classified for the study in the end in 3 groups counting 4 animals each. Pigs in the same group were operated with one of the above-mentioned techniques. It was found out that the gentlest method of all from the actual microcirculation level impact point of view is suture.
The drop in microcirculation intensity at the anastomotic site was to 59.7% maximum to the original values during 120 min after anastomosis construction. A more significant drop in microcirculation from the original values was in stapling and gluing to 39.6% and 34.5%, respectively. An increase in microcirculation intensity in glued anastomosis to 46.7% from the original values was observed. Macroscopically there was no disorder in healing in any animal recorded and microscopic examination showed satisfactory healing in all cases.
Conclusions
Partial ischemia of the large bowel tissues in the first 120 min after anastomosis construction is significantly higher in stapling and gluing. Function and biopsy results do not show significant differences in healing of the bowel whether the reconnection was performed by stapling, suture or glue.
Acknowledgment
The authors thank for supplementation of this experimental study by the Grant Agency of Charles University, Prague – project No. 187515.
References
1. Hardy KJ. A view of the development of intestinal suture. Part I. From legend to practice.
Aust NZ Surg 1990;60:299-304.
2. Hardy KJ. A view of the development of intestinal suture. Part II. From legend to practice.
Aust NZ Surg 1990;60:377-84.
3. Hardy KJ. Non-suture anastomosis: the historical development. Aust NZ Surg 1990;60:625-33.
4. Löffler T, Seiler CHM, Rossion I et al. Hand-suture versus stapling for closure of loop ileostomy: HASTA-Trial: a study rationale and design for a randomized controlled trial.
Trials 2011;12:1-10.
5. Páral J, Subrt Z, Lochman P et al. Suture-free anastomosis of the colon. Experimental comparison of two cyanoacrylate adhesives. J Gastrointest Surg 2011;15:451-9.
FACIAL NERVE PALSY AND TASTE DISTURBANCES Hana Faitlová, MUDr.
faitlovh@lfhk.cuni.cz
Department of Otorhinolaryngology and Head and Neck Surgery, Pardubice Hospital Department of Pathological Physiology, Faculty of Medicine in Hradec Králové, Charles
University in Prague
Co-authors: M. Bukvová, J. Vodička, H. Shejbalová Tutor: prof. MUDr. Miroslav Kuba, CSc.
Introduction
Etiology of an acute peripheral facial nerve palsy as a mononeuropathy is various (neurologic, infectious, parainfectious, traumatic, oncologic, vasomotoric, etc ) and in about one third of patients is uknown. Appellations "Bell's palsy", "idiopathic facial paralysis" and "e frigore"
are often used in these cases. There is a rising number of those caused by infection agents stated by new diagnostic methods. Anyway many infections remain still unrecognized, whereas diagnosis "e frigore" palsy (caused by vasospasm consequently cold temperature) is overused. [1,2]
Taste disturbance in patients with facial nerve palsy is usually investigated by simply query.
There are two kinds of taste dysfunctions in facial nerve palsy - regional hypo/ageusia (asymptomatic) and regional fantogeusia or parageusia (symptomatic). [3]
Aims
Having experience with asymptomatic unilateral taste loss in patient after middle ear surgery we were interested whether there is a taste loss in patients with facial nerve palsy and we also looked for possible relationship of gustatory dysfunction and etiology of the paresis.
Methods
We used Taste Strips test (Landis 2009 [4]) for investigation of taste function. Following qualities were tested: salty (NaCl), sweet (sucrose), sour (citric acid), bitter (quininehydrochloride). We enrolled 68 patients (35 males, 33 females) of age 6-89 years (mean 39, median 33 years). Patients were asked about the taste function. The cause of paresis was recorded. Patients were sorted out in two groups – “WITH” and “WITHOUT” taste disturbances. We compared the representation of palsy causes in each group.
Results
Impaired taste was detected in 81% (55) of patients, but clinical symptoms were reported by only 47% (32) patients. Etiology was idiopathic 53% (36), viral 29% (20), borreliosis 13%
(9), and injury 5% (3). In the group “WITH” impaired taste (N = 55) the causes were idiopathic 53% (29), viral 36% (20), borreliosis 9% (5), injury 2% (1). In the group
“WHITHOUT” taste disorder (N = 13) the causes ofwere idiopathic 54% (7), viral 0,
borreliosis 31% (4), injury 15% (2). Recurrent facial nerve palsy was observed in 7 patients, in all of these patients dysgeusia was found.
Discussion
Bell’s palsy is an acute mononeuropathy (peripheral lesion) of facial nerve. It was considered as a banal diagnosis with no necessity of other detailed examination [1]. The etiology is unknown and the theory of “e frigore” used to be generally accepted. Last two decades there is rising number of cases with confirmed infectious etiology as the diagnostic tools have been improving. [1,2] Several studies have demonstrated a link to herpes simplex virus in geniculate ganglion. [2] The facial nerve palsy should be considered as a symptom, but not as a diagnosis.
Part of these patients suffer from gustatory dysfunction that usually appears at onset of paresis or shortly in advance. Patients usually describe dysgeusia as a metallic taste or more often numbness of a half of a tongue ipsilateral to lesion. Last few years we have been focused on taste in patients after middle ear surgery and we found that some of them do have an unilateral taste lost being not aware of it. In literature this is explained as a “release of lateral inhibition phenomenon” [3]. We wondered if it could appear also in patients with facial nerve palsy.
Since chorda tympani with taste fibers leaves facial nerve above foramen stylomastoideum (with only a few exceptions), we suppose that in paresis “e frigore” should be no taste disturbance whereas infectious neuropathy (especially viral) should cause gustatory dysfunction.
Conclusions/Summary
Taste disturbance was found in 81% patients and only 47% patients were aware of it. Taste must be investigated not only by query (fantogeusia) but also examinated (asymptomatic regional hypo/ageusia). The Taste Strip Test (Landis 2009) is worthwhile as it enables to distinguish regional taste dysfunction.
All patients with evidence of viral cause of paresis and all patients with recurrent paresis had gustatory dysfunction. In patients having gustatory dysfuncion (either symptomatic or asymptomatic) infection or other more severe condition should be considered regardless anamnesis suggesting possible “e frigore” etiology.
References
[1] Bojar M.: Obrna lícního nervu. Cesk Slov Neurol N 2007; 70/103(6): 613-624.
[2] Hohman MH, Hadlock TA:Etiology, Diagnosis, and Management of Facial Palsy: 2000 Patients at a Facial Nerve Center Laryngoscope, 124:E283–E293, 2014
[3]Vodička J, Faitlová H: Poruchy čichu a chuti, Tobiáš, 2012, ISBN 978-80-7311-125-0 [4] Landis BN et al: “Taste Strips” – A rapid, lateralized, gustatory bed side identification test based on impregnated filter papers. Journal of neurology 2009;256(2):242-8.
Supported by grant SVV-2015-260179.
NATURALLY DERIVED SCAFFOLD FOR SKELETAL MUSCLE REGENERATION
Mgr. Hana Hrebíková hrebikh@lfhk.cuni.cz
Department of histology and embryology, Faculty of Medicine in Hradec Králové, Charles University in Prague
Co-authors: D. Díaz, P. Beznoska, A. Bezrouk, R. Pisal, J. Mokrý
Introduction: One of the major aims in regenerative medicine has been to developed solid organ and recent research works is focused on decellularization as an approach providing framework for building a new organ [1]. Decellularization mines from utilization of biomaterials and thus final biologic construct retains its complex geometry, including the functional aspects of native microvasculature [2]. Products of decellularization could find its use in clinical practise as demineralized bone matrix, skin grafts or biophrostetic heart valves [3,4,5]. The extracellular matrix (ECM), as a final result of decellularization, is
3D environment which can support adhesion, proliferation and differentiation of cells which are responding to signals from ECM and these signals can alter cells behaviour [6]. When properly processed to remove cellular components, ECM serves as a potent source of biomaterial.
Aim: The goal of this study is to find a suitable method for decellularizing skeletal muscle tissue, evaluate its effectiveness, analyse biocompatibility of the decellularized scaffolds with murine myoblasts and its transplantation into murine organism.
Methods: The protocol is based on the immersion of tissues in detergent solution in
combination with agitation including following steps: first step is disruption of cell membrane by osmotic shock which can improve penetration of subsequent detergent (1% SDS). This is followed by immersion with peracetic acid suitable for partial DNA removal which is subsequently assured by incubation with DNase. Scaffold is rinsed with sterile PBS buffer which is also used between every step for washing the scaffold. Decellularization efficiency was determined by histological staining (haematoxylin-eosin, Alcian blue, Sirius Red).
Morphological characteristics were examined by transmission electron microscopy (TEM).
The preservation of the ECM components was assessed by immunohistochemistry
(collagen IV, laminin). Preservation of collagen as a main component of ECM was assessed by biochemical method: hydroxyproline assay. Removal of cell nuclei was determined by quantification of DNA. Cytocompatibility was tested by seeding the muscle scaffolds with murine myoblasts for 12 days. Decellularized scaffold was transplanted into mouse and histologically analysed after 1 to 4 weeks (haematoxylin-eosin).
Results: Native (physiological) murine tibialis anterior muscle that underwent process of decellularization yielded a final translucent scaffold, which retained its shape and consistency.
Histological analysis proved the absence of any cell nuclei in paraffin-embedded sections counterstained with basic nuclear stain haematoxylin and revealed absence of sarcoplasmic
structures and cross-striated myofibrils while general skeletal muscle microarchitecture with ECM stromal components remained well preserved. TEM confirmed conclusions from histology: absence of cytoplazmatic and nuclear components, presence of ECM structures as collagen fibrils disclosing its striation. ECM glycoprotein, laminin, was also well preserved as documented by immunofluorescence analysis, which confirmed the glycoprotein was
localized to the basal lamina. Western blot also revealed bands at 250 kDa in the muscle scaffold corresponding to a full size of laminin. Quantification of residual DNA content after decellularization was significantly decreased from 113 ng DNA/mg of tissue in native muscle tissue sample compared to the 38 ng DNA/mg of tissue in a decellularized sample. Collagen, as the most abundant protein in ECM was quantified with hydroxyproline assay and in a native muscle tissue was 9.5 mg per mg of tissue compared to treated samples where was 83 mg per mg. Collagen presence was assessed as well by specific staining, Sirius Red, and with immunohistochemistry to prove preservation of collagen IV which is important
component of basal lamina. Adhesion and viability of the scaffold was confirmed by re-seeding with C2C12 murine myoblasts and examined in paraffin-embedded sections at 12 days after cultivation. Histological examination of scaffolds implanted into the murine tibial anterior muscle confirmed its good integration with the muscle and rich colonization by recipient cells.
Conclusion: We succeeded in producing decellularized skeletal muscle tissue with well- preserved major components (basal lamina, collagen fibers and glycosaminoglycans) that fulfilled requirement for successful decellularization (elimination of cellular and nuclear components, reduction of DNA content and preservation of 3D architecture of ECM).
Biocompatibility of scaffold was proved with a successful in vitro recellularization technique and by implantation of decellularized scaffold into the mouse muscle, which resulted in rich cell infiltration.
References:
[1] Badylak SF, Taylor D, Uygun K. Whole-organ tissue engineering: Decellularization and recellularization of three –dimensional matrix scaffolds. Annu Rev Biomed Eng. 2011;13:27- 53
[2] Qazi TH, Mooney DJ, Pumberger M, Geissler S, Duda GN. Biomaterials based strategies for skeletal muscle tissue engineering: Existing technologies and future trends.Biomaterials 2015;53:502-521
[3] Pacaccio DJ, Stern SF. demineralized bone matrix:basic science and clinical application.
Clin Podiatr Med Surg. 2005;22(4)599-606
[4] Hrebikova H, Diaz D, Mokry J. chemical decellularization: a promising approach for preparation of extracellular matrix. Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub. 2015;159(1):12-7
[5] Gilbert TW. Strategies for tissue and organ decellularization. J Cell Biochem.
2012;113(7):2217-2222
[6] Nelson CM, Bissell MJ. Of extracellular matrix, scaffolds and signalling: Tissue architecture regulates development, homeostasis and cancer. Annu Rev Cell Dev Biol.
2006;22:287-309
ANTIMICROBIAL FOOD ADDITIVES INFLUENCE THE COMPOSITION OF HUMAN GUT MICROBIOTA
Lucia Hrnčířová, MD SAMAL8A1@lfhk.cuni.cz
Department of clinical immunology and allergology, Faculty of Medicine in Hradec Kralove, Charles University in Prague
Co-authors: T. Hrncir, E. Trckova, N. Gabrielova, J. Krejsek Tutor: Prof. RNDr. J. Krejsek, CSc.
Introduction
There is an accumulating evidence that gut microbiota play an important role in the induction and maintenance of various allergic, autoimmune, metabolic diseases and cancer [1,2,3]. The ratio of beneficial (anti-inflammatory) versus harmful (pro-inflammatory) microbiota seems to play a central role [4]. The composition of gut microbiota is influenced by genetic and environmental factors. The single most important environmental factor is diet [5].
A preservative is naturally occurring or synthetically produced substance that is added to food to prevent microbial growth or undesirable chemical changes. Preservative food additives can be used alone or in combination with other methods of food preservation. Preservatives may be antimicrobial preservatives, which inhibit the growth of bacteria or fungi, including mold, or antioxidants, which inhibit the oxidation of food constituents. Common antimicrobial additives include sorbic acid and its salts, benzoic acid and its salts, potassium sorbate, calcium propionate, sodium nitrite, sulfites and disodium EDTA.
Aims
The aim of the project is to evaluate whether and to which extent commonly used food additives modify the composition of gut microbiota.
In vitro part: to test inhibitory effect of widely used antimicrobial food additives on single bacteria isolated from human gut microbiota
In vivo part: to test the influence of antimicrobial food additives on the composition of human gut microbiota (experiments with germ-free mice colonized with human gut microbiota) Methods
We have isolated six aerobic bacteria from human fecal sample. We have grown aerobic bacteria in a tryptic soya broth to OD600 1.0 and incubated them, for 24 hours at concentration of 10^3 bacteria per well, with antimicrobial food additives, namely sodium benzoate, sodium nitrite, potassium sorbate, and sodium sulfite, at different concentrations. Then minimal inhibitory concentrations (MIC) and fractional inhibitory concentration (FIC) were determined using spectrofotometry.
To address in vivo experiments we colonized germ-free C57BL/6 mice with the microbial community sample from a healthy adult human. Experimental mice were treated with selected food additives supplemented in drinking water. The composition of gut microbiota was identified using bacterial community analysis (QIIME) of sequenced genomic DNA from fecal samples.
Results
The MIC of sodium benzoate, potassium sorbate and sodium nitrite is different for particular AMFAs and isolated aerobic human gut bacteria, MIC of sodium sulfite is the same for each of them and much higher concentrations are needed to inhibit growth. The FIC measurements show that sodium
benzoate and sodium nitrite have a synergistic effect on Enterobacter, one strain of Klebsiella and one strain of E.coli. Sodium benzoate and potassium sorbate have a synergic effect on Enterobacter, Klebsiella, and one strain of E.coli. Sodium nitrite and potassium sorbate have a synergistic effect on Enterobacter, Klebsiella, and one strain of E.coli. The effect of sodium sulfite was significantly different from other AMFAs.
When tested the effect of the presence of AMFA in diet in vivo, we have found that low AMFA concentrations decrease the diversity of human gut microbiota, specifically the Actinobacteria and Verrucomicrobia phyllum disappear, and increase the representation of the Proteobacteria Phyllum, high AMFA concentrations in diet entirely disrupt the gut microbiota ecosystem with the Proteobacteria phyllum overgrowth.
Discussion
We have tested the inhibitory effect of four common AMFAs on the growth of culturable gut bacteria.
We have found that this effect is different for each bacteria and AMFA and can be synergistic in some conditions, so we hypothesize, that the presence of many different AMFAs in diet can influence the growth of single gut bacteria and contribute to dysbiosis, albeit most of gut bacteria are anaerobic and unculturable. The effect of sodium sulfite was different from the others, it seems that it can react with others AMFAs to prevent its acting.
The in vivo experiments were performed with human gut microbiota from one donor, so we can not exclude the resulting effect of AMFAs in diet on the composition of gut microbiota is universal.
Conclusions/Summary
To study the effect of preservatives on the composition of human gut microbiota we performed in vitro studies and for in vivo experiments we exploit a humanized mice model. In vitro, AMFAs inhibit the growth of human gut bacteria in a different manner and in some concentrations, the growth inhibitory effect is synergistic.
Our preliminary data from microbiome profiling of additive-treated mice colonized with human gut microbiota show that antimicrobial food additives have the capacity to alter the composition of gut microbiota.
We have found, that the AMFAs present in diet have the potential to influence the composition of human gut microbiota. Further work is need to ascertain the impact of changed microbiota on the function of immune system and the development of disease.
References
[1] Belkaid Y, Hand TW, Role of the microbiota in immunity and inflammation. Cell. 2014 Mar 27;157(1):121-41.
[2] Shen J, Obin MS, Zhao L, The gut microbiota, obesity and insulin resistance. Mol Aspects Med.
2013 Feb;34(1):39-58
[3] Louis P, Hold GL, Flint HJ. The gut microbiota, bacterial metabolites and colorectal cancer. Nat Rev Microbiol. 2014 Oct;12(10):661-72
[4] Sommer F, Bäckhed F. The gut microbiota--masters of host development and physiology. Nat Rev Microbiol. 2013 Apr;11(4):227-38.
[5] Thorburn AN, Macia L, Mackay ChR Diet, Metabolites, and “Western-Lifestyle”Inflammatory Diseases. Immunity. 2014 Jun 19;40(6):833-42
This work was supported by Charles University in Prague, Faculty of Medicine in Hradec Kralové, Czech Republic, project “PRVOUK” P37/10 and by grant from the Grant Agency of Charles University in Prague (No. 906613).
Neonatal outcomes in subgroups of women with preterm prelabor rupture of membranes before 34 weeks
Daniel Lesko, MD daniel.lesko@email.cz
Faculty of Medicine Hradec Kralove, Department of Obstetrics and Gynecology, Charles University in Prague, Hradec Kralove, Czech Republic
Co-authors: Martin Stepan, Teresa Cobo, Jan Maly, Martina Navratilova, Ivana Musilova,Helena Hornychova, Bo Jacobsson, and Marian Kacerovsky
Tutor: doc.MUDr. Jiri Spacek Ph.D.,IFEPAG Abstrakt
Objective:
To evaluate the influence of microbial invasion of the amniotic cavity (MIAC) and
histological chorioamnionitis (HCA) on short-term neonatal outcome in women with pretermprelabor rupture of membranes before 34 weeks of gestation.
Methods:
A prospective observational cohort study including 122 pregnant women with
PPROM between 24+0 and 34+0. MIAC was defined as a positive PCR result for Ureaplasmaspecies, Mycoplasma hominis and Chlamydia trachomatis and/or positive PCR result for the16S rRNA gene in the amniotic fluid. HCA was defined according to the Salafia classification. Maternal and short-term neonatal outcomes were evaluated according to the presence or absence of MIAC and/or HCA.
Results:
The presence of both MIAC and HCA was observed in 36% (45/122) of women, HCA
alone in 34% (41/122) and MIAC in 5% (6/122). A significantly higher incidence of early onset sepsis was observed in newborns born from women with both MIAC and HCA [33% (15/45)]
compared with women with HCA alone [12% (5/41)] or MIAC alone [0% (0/6)] or women without MIAC or HCA detected [0% (0/30); pĽ0.001].
Conclusions:
The presence of both MIAC and HCA increases the risk of early onset sepsis in
pregnancies complicated by preterm prelabor rupture of membranes before 34 weeks of gestation.
References:
1. Parry S, Strauss JF. Premature rupture of the fetal membranes. N Engl J Med 1998;338:663–70.
2. Johanzon M, Odesjo H, Jacobsson B, et al. Extreme preterm birth: onset of delivery and its effect on infant survival and morbidity. Obstet Gynecol 2008;111:42–50.
3. Morken NH, Kallen K, Hagberg H, Jacobsson B. Preterm birth in Sweden 1973-2001: rate, subgroups, and effect of changing patterns in multiple births, maternal age, and smoking. Acta Obstet Gynecol Scand 2005;84:558–65.
4. Suppiej A, Franzoi M, Vedovato S, et al. Neurodevelopmental outcome in preterm histological
chorioamnionitis. Early Hum Dev 2009;85:187–9.
5. Been JV, Zimmermann LJI. Histological chorioamnionitis and respiratory outcome in preterm infants. Arch Dis Child Fetal Neonatal Ed 2009;94:F218–25.
6. Buhimschi CS, Buhimschi IA, Abdel-Razeq S, et al. Proteomic biomarkers of intra-amniotic inflammation: relationship with funisitis and early-onset sepsis in the premature neonate. Pediatr Res 2007;61:318–24.
7. Shim SS, Romero R, Hong JS, et al. Clinical significance of intraamniotic inflammation in patients with preterm premature rupture of membranes. Am J Obstet Gynecol 2004;191:1339–45.
8. Cobo T, Palacio M, Martinez-Terron M, et al. Clinical and inflammatory markers in amniotic fluid as predictors of adverse outcomes in preterm premature rupture of membranes. Am J Obstet Gynecol 2011;205:126 e121–8.
9. Russell P. Inflammatory lesions of the human placenta. I. Clinical
significance of acute chorioamnionitis. Am J Diagn Gynecol Obstet 1979;1:1.
10. Tsiartas P, Kacerovsky M, Musilova I, et al. The association
between histological chorioamnionitis, funisitis and neonatal outcome in women with preterm prelabor rupture of membranes. J Matern Fetal Neonatal Med 2013;26:1332–6.
11. Cobo T, Kacerovsky M, Palacio M, et al. Intra-amniotic inflammatory response in subgroups of women with preterm prelabor rupture of the membranes. PLoS One 2012;7:e43677.
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795–801.
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response in women with proteomic biomarkers characteristic of intra-amniotic inflammation and preterm birth. Bjog 2009;116: 257–67.
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16. The Czech Society of Obstetrics and Gynecology. 2007. Clinical guidelines in obstetrics.
Available in Czech language from: http:// wwwperinatologiecz/dokumenty.Q2
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22. Kacerovsky M, Musilova I, Hornychova H, et al. Bedside assessment of amniotic fluid interleukin-6 in preterm prelabor rupture of membranes. Am J Obstet Gynecol 2014;211:385e381–9.
Influence of right ventricular pacing on right ventricular systolic function Background and aim of the study:
Even though right ventricle pacing is irreplaceable therapeutic modality, the question of it´s influence on myocardial contractility remains remains unresolved.
Aim of this prospective study is to inquire, whether right ventricle pacing worsens the right ventricle systolic function and, if need be, which position of the right ventricle pacing lead is more favorable with regard to the right ventricle systolic function.
We have also tested the ability of real time 3D transthoracic echocardiography (TTE) to verify the position of the pacing lead.
Methods:
30 patients were included in the study in the period since March 2013 to March 2015. This population was randomly divided in to two groups: with apical and septal position of the lead (11 and 19 pts.). Apical position was chosen by the implanting physician in case of provocation of electric instability by the septal position or in case of unsatisfactory electric parameters in the septal position of the lead. To verify the lead position, real-time 3D echo on Philips IE33 machine was used, besides the more usual methods (fluoroscopy and ECG).
For evaluation of the systolic function of the right ventricle we have used tricuspid annulus plain systolic excursions (TAPSE) and tricuspid annulus systolic velocity (TASV) parameters. Values under 16 mm of TAPSE and under 10 cm/s of TASV were considered decreased.
The percentage of ventricular pacing (VP) was red from the external programmers.
Used pacing modes were DDD, DDDR, VVI, VVIR, AAI-DDD and AAIR-DDDR.
The time of follow up was 2-26 months, average 8.1 months.
Demographic parameters of the population:
Population consists of 19 male and 11 female patients, aged 45-93 years, average age was 74 years. There were 11 apical and 19 septal lead positions. 6 pacemakers were one chamber and the rest (24) were two chamber devices. The average percentage of ventricular pacing (VP) was 62%. 14 patients were dependend on the pacemaker with VP of 99% or more.
Statistical analysis:
In order to inquire, whether or not, there is a significant decrease of TAPSE and TASV values, in all patients with right ventricle pacing, we have used the pair t-test. We have studied the differences (d) in the pair couples of TAPSE and TASV 1 and 2 (first and second measured values) and we have tested the zero hypothesis H0, that the values of pair couples do not differ systematically in one direction. The normality of the selected samples was verified by the Chisq test of the good concurrence and also by the Kolmogorov-Smirnov test. The measured values of TAPSE: d=0.8±0,848, i.e. Confidence interval (CI) for d is (-0.05; 1.65), and for TASV d=0.483±0.694 i.e. CI for d is (-0,216;1,15).
Pair test showed, that there is not a significant difference – decrease of the values of both parameters – TAPSE p=0.0746 and TASV p=0,19055.
We have also studied influence of the lead position on the difference between first and second measurement: The difference between pair values of TAPSE and TASV in two independent groups of patients, that is, group with apical (n=11) and group with non-apical (n=19) lead position. We have used non-parametrical Mann-Whitney test. H0: Lead position does not influence the differences of the values in the given two groups – H0 is not rejected on the level of significance 5% with p=0.2 for TAPSE and p=0.11 for TASV.
Results:
1. Pair test showed, that there is not a significant difference – decrease of the values of both parameters – TAPSE and TASV in all patients with right ventricle pacing and
2. non-parametrical Mann-Whitney test showed, that H0 (Lead position does not influence the differences of the values in the given two groups) is not rejected on the level of significance 5%, p=0.2 for TAPSE and p=0.11 for TASV.
3. There was not a single patient, in whom there would be a decline of any studied parameter under the limit of normal values, given by the American Society of Echocardiography (1).
4. In all patients included in the study, it was possible to visualize the ventricular lead with the use of transthoracic echo. Since the real time 3D echo is able to show a longer part of the lead at the same time than 2D TTE, it provided either better information than 2D or at least the same. Real-time 3D TTE often provides a better visualization of the lead insertion in to the myocardium.
Discussion:
The right ventricle pacing has been used for more than 5 decades and this method significantly prolongs and improves the quality of life of the population. Yet, one of the possible side effects is worsening of the systolic function of the left ventricle (LV). This question has been discussed for more than 20 years. 14 randomized trials were performed, of which, some suggest a negative influence of the apical right ventricular pacing on the LVEF. Strongest evidence of this has been brought for patients who had systolic dysfunction of the LV already before the implantation.
In a systematic review and meta-analysis of 14 RCTs for a total of 754 patients, those randomized to RV non-apical pacing had greater LVEF at the end of follow-up especially those with a baseline LVEF <45%
and with a follow-up length >12 months. No significant difference was observed in RCTs of patients whose baseline LVEF was preserved. Results were inconclusive with respect to exercise capacity, functional class, quality of life and survival. This Task Force is unable to give definite recommendations until the results of larger trials become available. (2)
Pacing of the right ventricle myocardium, other than the His-Purkinje conduction system can cause a decrease in stroke volume and abnormal relaxation of the LV (3).
Some studies demonstrate, that long term RV pacing, causes LV remodeling with asymetric hypertrophy and dilatation, mitral regurgitation, decrease in myocardial perfusion and decrease in ejection fraction (4) Worsening of the systolic function of the right ventricle plays a major role in the morbidity and mortality of patients with cardiopulmonary disease. Despite that, there has been to date, to the best of our knowledge, only one work, studying the influence of the RV pacing on the RV function and this was a retrospective analysis of 96 pts. Prevalence of the RV dysfunction in this population was 4%, but, one of the limitations of this study is, that the RV function was not evaluated before the implantation (4).
Evaluation of the RV function: Even though RV has more complex anatomy and kinetics, parameters TAPSE and TASV are easy to obtain, and have a very good reliability and reproducibility. The use of TAPSE as a parameter, evaluating the right ventricle systolic function has been repeatedly verified by a series of studies correlating TAPSE with MRI or radionuclide ventriculography. Values of 16 mm and more are, according to the current guidelines, considered normal. (1). Another parameter, assessing the quality contractions of longitudinal fibres is St or TASV (tricuspid annular systolic velocity), obtained by the pulse-tissue Doppler ultrasound of the lateral margin of the tricuspid annulus. Again, the correlation of TASV with right ventricle ejection fraction has been confirmed by multiple radionuclide and MR imaging studies. In both of these parameters, a strong prognostic information - like in patients with heart failure or patients with pulmonary artery hypertension - has been proved. That is why these parameters are recommended for the routine evaluation of the right ventricle systolic function.
Studies, focusing on the negative impact of right ventricular pacing on the systolic function of the heart, were always evaluating LVEF, which parameter has also it´s limitations, like imperfect reproducibility, especially in the case of suboptimal image quality.
The author is aware of the fact, that parameters of RV function are not applicable to the assessment of LV function, but, even though left and right ventricles are anatomically two different parts of one organ,
histologically they constitute integrity of one syncytium and thus, should there be a strong and clear evidence, that RV function is really not worsened by the RV pacing from any site, it would constitute certain questioning of the concept of negative influence of the RV pacing on the myocardial function as a whole.
Conclusion: Based on these data, it seems unlikely, that right ventricle pacing influences the right ventricular systolic function.
At the same time we have confirmed, that TTE is a very good method for determining the ventricular lead position by it´s direct visualization.
Literature:
1. Rudski LG, Lai WW, Afilalo J, et al. Guidelines for the Echocardiographic Assessment of the Right Heart in Adults: A Report from the American Society of Echocardiography Endorsed by the European Association of Echocardiography, a registered branch of the European Society of Cardiology and the Canadian Society of Echocardiography. J Am Soc Echocardiogr 2010; 23: 685-713.)
2. 2013 ESC Guidelines on cardiac pacing and cardiac resynchronization therapy. European Heart Journal (2013) 34, 2281–2329 doi:10.1093/eurheartj/eht150. (2)Shimony A, Eisenberg MJ, Filion KB, Amit G, Beneficial effect of right ventricular pacing non-apical pacing vs. apical pacing: a systematic review and meta-analysis of randomized-controlled trials. Europace 2012; 14: 81-91.)
3. Lee MA, Dae MW, Griffin JC, et al. Effects of long-term right ventricular apical pacing on left ventricular perfusion, innervation, function and histology. J Am Coll Caardiol 1994; 24: 225-232
4. Tse HF, Lau CP. Long –term effect of right ventricular pacing on myocardial perfusion and function. J Am Coll Cardiol 1997; 29: 744-9, Tantengco MV, Thomas RL, Karpawich PP. Left ventricular dysfunction after long-term right ventricular apical pacing in the young. J Am Coll Cardiol 2001; 37: 2093- 100)
5. Pornwalee Porapakkham et. al. Impact of Right Ventricular Pacing on Right ventricular funcion, J Med Assoc Thai 2012; 95 (Suppl. 8): S44-S50
TOXIC EFFECT OF 3-BROMOPYRUVATE ON RAT HEPATOCYTES IN VITRO Ondřej Sobotka
sobotkao@lfhk.cuni.cz
Department of Physiology, Faculty of Medicine in Hradec Kralove, Charles University in Prague Co-authors: R. Endlicher, Z. Drahota, O. Kučera, D. Rychtrmoc, M. Raad, K. Hakeem, Z. Cervinkova
Tutor: Prof. MUDr. Zuzana Cervinkova, CSc.
Introduction
3-bromopyruvate (3-BrPyr) is a well-known alkylating agent with documented anticancer effects [1]. 3-BrPyr induced growth arrest of implanted tumors in vivo and caused necrosis and apoptosis of different transformed cell lines in vitro [2]. The inhibition of several intracellular enzymes influencing the homeostasis of the cell was described [3].
Aims
The main goal of this study was to assess the effect of 3-BrPyr on primary rat hepatocytes exposed for up to 20 hours. We focused on a viability, functional capacity, intracellular ROS production, type of cell death, morphological state of the culture and possible mitochondrial damage. The importance of this study is potentiated by the first use of 3-BrPyr in human cancer treatment [4].
Methods
WST-1 test was used to assess the activity of intracellular dehydrogenases. The damage of cell membrane was evaluated by the measurement of lactate dehydrogenase (LDH) activity in extracellular medium and by calculated LDH leakage. The functional state of cultured hepatocytes was estimated by measurement of total albumin production. The concentration of albumin was measured by ELISA method. ROS production was evaluated using DCFDA fluorescent probe. The presence of apoptotic cell death was assessed by the caspase 3 activity measurement. Phase contrast microscopy was used to evaluate cell morphology and fluorescent probes JC-1 and DAPI enabled to visualize the mitochondrial membrane potential (MMP) and to illustrate apoptotic changes respectively. After short term 3-BrPyr exposure, Oroboros O2k respirometry and Safranin O fluorescent probe were used to measure direct effect of 3-BrPyr on stimulated mitochondrial oxygen consumption and MMP respectively.
The data were obtained from at least three independent hepatocyte isolations and statistical evaluation was performed using GraphPad Prism software.
Results and Discussion
We demonstrated toxic effect of 3-BrPyr on non-cancer rat hepatocytes in vitro. 3-BrPyr caused damage to the morphological structure and attenuated viability and functional capacity of cultured hepatocytes. Both the diminished activity of cellular dehydrogenases and greater plasmatic membrane damage exhibited dose- and time-dependent response relationship.
Albumin production after 20 hours of exposure to 3-BrPyr was significantly decreased from concentration 150 µmol/l of 3-BrPyr in culture medium (p<0.001). The cellular damage was accompanied with increased ROS production. We found significantly increased ROS production after one hour of incubation with 3-BrPyr at concentrations ≥100 µM (p<0.01).
We proved higher activity of caspase 3 after 20 hours of incubation at concentrations 150 µM and 200 µM of 3-BrPyr, however at concentration 300 µM the activity of caspase 3 was under the detection limit. The finding was confirmed by the DAPI fluorescence staining. We observed chromatin fragmentations and condensations, which are changes typical for programmed type of cell death. Fluorescence staining JC-1 showed decrease in MMP.
Our results indicate possible connection with mitochondrial dysfunction. To differentiate if the mitochondrial dysfunction is the cause or consequence we further investigated the direct effect of 3-BrPyr on mitochondrial properties. 3-BrPyr induced decline in MMP for glutamate and malate stimulated respiration from concentrations ≥ 20 µM (incubation time 10 minutes;
p<0.001). The respiratory rate for complex I initiated mitochondrial respiration was similarly reduced. However succinate stimulated respiration and MMP was even more sensitive starting at concentration 10 µM of 3-BrPyr (p<0.001) in suspensions of permeabilized hepatocytes.
This effect of 3-BrPyr on electron transport chain was confirmed by the measurements on isolated rat liver mitochondria. The results on isolated mitochondria show stronger 3-BrPyr effect, confirming our assumption that mitochondria play significant role in 3-BrPyr effect on non-cancer rat hepatocytes.
We succeeded to provide the evidence of 3-BrPyr toxic action on primary hepatocytes in vitro. This toxic effect of 3-BrPyr on non-cancer tissue was pronounced even in concentrations comparable to doses used in experiments with HepG2 cell line [5]. We want to point out the fact that the toxic effect is connected with mitochondrial dysfunction and initiation of the apoptotic pathway. According to our results low doses of 3-BrPyr induce apoptosis and higher doses lead directly to necrotic damage. The results of this study need to be reconsidered in upcoming in vivo experiments or in future clinical trials.
Acknowledgements
This study was funded by PRVOUK P37/02 References
1. Pedersen, P.L., 3-Bromopyruvate (3BP) a fast acting, promising, powerful, specific, and effective "small molecule" anti- cancer agent taken from labside to bedside: introduction to a special issue. J Bioenerg Biomembr, 2012. 44(1): p. 1-6.
2. Ko, Y.H., et al., Advanced cancers: eradication in all cases using 3-bromopyruvate therapy to deplete ATP. Biochem Biophys Res Commun, 2004. 324(1): p. 269-75.
3. Shoshan, M.C., 3-Bromopyruvate: targets and outcomes. J Bioenerg Biomembr, 2012. 44(1): p. 7-15.
4. Ko, Y.H., et al., A translational study "case report" on the small molecule "energy blocker" 3-bromopyruvate (3BP) as a potent anticancer agent: from bench side to bedside. J Bioenerg Biomembr, 2012. 44(1): p. 163-70.
5. Pereira da Silva, A.P., et al., Inhibition of energy-producing pathways of HepG2 cells by 3-bromopyruvate. Biochem J, 2009. 417(3): p. 717-26.
1
Possibilities of lesional vas deferens repair in the experiment
Radek Štichhauer, MD stiradek@centrum.cz
Academic Department of Surgery, Faculty of Medicine, Charles University, Hradec Králové Co-authors: J. Koudelka, K. Petkov, A. Ryška, M. Kaška
Tutor: Assoc. Prof. Dr. Milan Kaška, MD, PhD Introduction
Reparation of inguinal hernia represents one of the most frequent general surgical operations both in adults and children. Some literature references regarding this theme speak about the fact that the tissues of resected hernia sacs can contain the tissue structures of the vas deferens in 0.1 – 1.7% in some analysed groups of the youngest children. This finding probably seems to be one of the most frequently occurring iatrogenic injuries which can happen in operations performed by paediatric surgeons. Surgical reparation of the lesional vas deferens in adults have been described in details, but recommendation for the appropriate therapeutic method for this injury in newborns and in a very young children group is missing. Microsurgery is not commonly performed at paediatric surgery departments for limitations in special apparatuses and special surgical devices equipment and in a surgeon’s experience. The main idea of the present experimental study is a creation of some useful algorithm for injured vas deferens reparation during inguinal hernia surgery under the conditions of our basic paediatric surgery departments with the use of magnifying glasses only.
Aims
The aim of this experimental study with a small laboratory animal is to perform and analyze the possibilities of new reparation methods for the lesional vas deferens in a standard paediatric surgery department without the use of a microscope.
Hypotheses
Injury of the small child vas deferens after its contusion by a surgical instrument can cause its substantial morphologic and functional changes. Methods of vas deferens dissection and its sequent surgical reparation with the application of various techniques and materials can lead to the finding of their best combination recommendable for a daily clinical practice use.
Materials and methods
A pilot prospective experimental study was performed on the rat, Wistar strain, body weight 350-450 g, whose anatomical condition of the vas deferens is very similar to those in very small children. 36 animals were included into the study and they were divided in six subgroups according to the method of the vas deferens injury (contusion or transection) and its consequent reparation. The subgroups of animals according to the method of injury and sequent reparation method: 1. contusion of the vas deferens by pressing in a pean for 2 sec, 2.
anastomosis of the vas deferens by single absorbable stitches (Vicryl R 8/0), 3. jointing of both ends each to other with the help of an intraluminally lead fibre of absorbable sewing material (PDS 8/0) knotted externally, 4. = 3. jointing with a non-absorbable fibre of sewing material (Prolen 7/0) knotted externally, 5. anastomosis by absorbable sewing material (Vicryl R 8/0) with an intraluminally situated fibre of absorbable sewing material (PDS 7/0), 6. anastomosis by absorbable sewing material (Vicryl R. 8/0) with an intraluminally situated fibre of non-absorbable sewing material (Prolen 7/0). All above mentioned operations were performed in general anaesthesia (ketamin + xylazin solution intraperitoneally) using minor surgical instruments and with the help of magnifying glasses (enlargement 4x). The vas deferens was checked 3 months after the primary operation. An operated both other side were resected in a length of 20 mm with a line of artificial injury in the central part. These resected
2 parts of the vas deferens were examined in function by the flow rate of methylene blue solution (mL/min). The pathologist performed morphologic evaluation of the resected vas deferens using histology with HE-staining. Findings on the injured vas deferens were compared with those on the second side without surgery.
Results
Surgery/reparation
Mean stream rate in operated VD
(mL / min.)
Mean stream rate in non-operated VD
(mL / min.)
Histologic findings
Histologic findings
Contusion/non 17.885 17.950 Minimal
changes
normal cut off/anastomosis 9.032 17.875 Tight stenoses
or dehiscence
normal cut off/ jointing
absorbable stitch
13.173 16.313 Small
changes
normal cut off/jointing
non-absorb. stitch
4,004 13.328 Tight stenoses
or dehiscence
normal cut off/anastomosis
+absorb. fibre
10.402 17.901 Stenoses
or dehiscence
normal
cut off/anastomosis + non-absorb. fibre
6.879 17.827 Tight stenoses
or dehiscence
normal
VD - vas deferens, anastomosis-suture by Vicryl 8/0 Conclusions
We found a normal liquid flow rate through the resected part of the vas deferens and morphological conditions in this subgroup with the contused vas deferens after 3 months. The best results in liquid flow rate through the transected and repaired vas deferens were evaluated in the subgroup with reparation performed by jointing the transected vas deferens with the help of absorbable sewing material (PDS 7/0) situated intraluminally and knoted out of the vas deferens. Similar results were found in the subgroup anastomosis performed with absorbable stitch (Vicryl - 8/0) and with intraluminally situated absorbable stitch (Vicryl 7/0) Examination of healed anastomoses carried out with absorbable sewing material 8/0 without an intraluminally situated stitch fibre as “a leader” and with “a leader” from non-absorbable sewing material showed the substantially worst results.
References
1. Ozen IO, Bagbanci B, Demirtola A, Karabulut Rozen O et al. A novel technique for vas deferens transection repairs. Pediatr Surg Int 2006; 22:815-19.
2. Steigman CK, Sotelo-Avila C, Weber TR. The incidence of spermatic cord structures in inguinal hernia sacs from male children. Am J Surg Pathol 1999; 23:880-5.
3. Sheynkin YR, Hendin BN, Schlegel PN, Goldstein M. Microsurgical repair of iatrogenic injury to the vas deferens. J Urol 1998; 159:139-41.
4. Vrijhof EJ, deBruine A, Lycklama AA, Koole LH. A polymeric mini-stent designed to facilitate the vasectomy reversal operation. A rabbit model study. Biomaterials 2004;
25:729-34.
Acknowledgments
The authors thank for support of this study the Grant Agency UK, Prague, No. 160315.
MATERNAL SERUM C-REACTIVE PROTEIN IN WOMEN WITH PRETERM PRELABOR RUPTURE OF MEMBRANES
Martin Štěpán, M.D.
mstepan.hk@post.cz
Department of Obstetrics and Gynecology, Charles University in Prague, Faculty of Medicine Hradec Kralove, Czech Republic
Co-authors: Teresa Cobo, Ivana Musilová, Helena Hornychová, Bo Jacobsson, Marian Kacerovský
Tutor: Assoc. Prof. Marian Kacerovský, M.D., PhD.
Abstract
Objective: To evaluate maternal C-reactive protein (CRP) as a predictor of both
microbial invasion of the amniotic cavity (MIAC) and histological chorioamnionitis (HCA) in women with preterm prelabor rupture of the membranes (PPROM) before and after 32 weeks of gestation.
Methods: Prospective observational cohort study including 386 women. Maternal serum CRP concentrations were evaluated and amniotic fluid samples were obtained through transabdominal amniocentesis at the time of admission. After delivery, the placenta was subjected to histopathological examination. MIAC was defined based on a positive PCR for Ureaplasma species, Mycoplasma hominis and Chlamydia trachomatis and/or through positive 16S rRNA gene amplification. HCA was defined according to the Salafia classification.
Results: The maternal CRP was significantly higher in women with both MIAC and HCA (median 9.0 mg/l) than in women with HCA alone (median 6.9 mg/l), with MIAC alone (median 7.4 mg/l) and without MIAC or HCA (median 4.5 mg/l);
p<0.0001. CRP showed a weak prediction of the occurrence of both MIAC and HCA before and after 32 weeks of gestation. Only the 95th percentile of CRP and only in PPROM below 32 weeks showed a false-positive rate of 1%, a positive predictive value of 90% and a positive Likelihood ratio of 13.2 to predict both MIAC and HCA.
However, the low sensitivity of 15% limits the clinical utility of this detection.
Conclusion: CRP poorly predicts the occurrence of both MIAC and HCA, even at early gestational ages.
