The organization of an educational program for specialists in clinical chemistry by the Greek Society of Clinical Chemistry-Clinical Biochemistry.

In Greece, there is no officially organized training in clinical chemistry for scientists. The Greek Society of Clinical Chemistry-Clinical Biochemistry decided to organize an intensive educational program of 18 seminars on clinical chemistry content as it is described in the EC4 Syllabus. The duration of each seminar was about 6 hours and consisted of 6 to 9 lectures. At the end of each seminar there was a voluntary written examination, comprised of 24 multiple choice questions. Successful completion of the Educational program was leading to a Certificate of Competence. Two cycles of the 18 seminars were performed: 1st cycle from October 2003 to December 2005 and 2nd cycle from March 2005 to October 2007. One hundred eighty nine colleagues was the mean attendance per seminar for the seminars of the 1st cycle and 38 colleagues for the seminars of the 2nd cycle. The mean participation to the examination for each seminar was almost 80% for the 1st cycle and 68% for the 2nd cycle. More than 80% of the participants performed Good or Very good in the examination in both cycles. It is estimated that more than 40% of the scientists who practice Clinical Chemistry in Greece, participated to this educational activity. This program is now provided as an e-learning application, and it is open for all scientists who want to follow the discipline of clinical chemistry.

The Greek way to the Register: the establishment and operation of the Register for Clinical Chemists-Clinical Biochemists in Greece.

In Greece, there is no officially organized training in clinical chemistry for scientists. The Greek Society of Clinical Chemistry-Clinical Biochemistry (GSCC-CB), following the encouragement of the EC4/RC decided to organize a voluntary Register for specialists in clinical chemistry. The following criteria for registration were defined: 1) University degree in Chemistry, Biochemistry, Biology, Medicine, Pharmacy or other relevant subject. 2) A total of 9 years of university studies and postgraduate specialization in clinical chemistry-clinical biochemistry. 3) A minimum of 4 years of postgraduate specialization in clinical chemistry-clinical biochemistry on the job. 4) The candidate must be practicing clinical chemistry-clinical biochemistry in a laboratory in a medical environment in Greece. The postgraduate specialization in clinical chemistry-clinical biochemistry includes the laboratory training and the theoretical education. The laboratory training is organized by the GSCC-CB according to the Professional Training Dossier. The theoretical education was organized in a series of 18 "Seminars" which was the content of the "Educational program" of the GSCC-CB. Successful completion of the Educational program leads to a Certificate of Competence. The Greek Register has gained equivalence with the EC4 Register and it has 218 members, more than 80 of whom are European clinical chemists.

Immune response parameters during labor and early neonatal life.

AIM: Selected cytokines, associated with Th1 and Th2 immune response and inflammation, were studied in order to evaluate the relation between their release into maternal and neonatal circulation, during labour, and after birth, in comparison with those in adults. MATERIALS AND METHODS: Cytokine concentrations were determined by very sensitive immunoassays, in maternal serum (MS), umbilical cord (UC), neonatal serum, the 1st (1N) and 5th (5N) day postpartum and in adult controls. RESULTS: Both IL-2 and IL-4 cytokine concentrations in UC were markedly elevated, compared to adult and MS ones. IL-2 decreased significantly in 5N, while IL-4 remained unchanged. IFN-gamma UC values were significantly lower than those in adults and MS, increasing significantly in 5N. Neonatal serum sIL-2R and sIL-4R were markedly higher than those in adults and MS. IL-1beta, IL-6, sIL-6R, sTNFRI and sTNFRII concentrations in MS and all with TNF-alpha in neonatal serum were significantly higher than in adults. IFN-gamma, IL-1beta, IL-6, TNF-alpha, IL-2R, IL-4R concentrations in MS, 1N and 5N were dependent on the mode of delivery. CONCLUSION: The results of this comparative study are indicative for a meaningful role for the studied cytokines and their receptors in: i) the development of neonatal immune system, ii) the regulation of immune response during labour and early life, and iii) the initiation of the processes of labour.

Tumor markers in biological fluids associated with pregnancy.

Proteins that are expressed by both malignant and healthy fetal tissues are recognized as oncofetal. These antigens are associated with cell proliferation and differentiation and are produced in high concentrations in pregnancy and malignancy. Their biological role in malignancy is the suppression of the host's immune system, while in pregnancy they affect the maternal immune response, generating maternal tolerance toward the embryo. This review describes the levels of alpha-fetoprotein (AFP), human chorionic gonadotropin (hCG), carcinoembryonic antigen (CEA), cancer antigen 125 (CA 125), squamous cell carcinoma antigen (SCC), cancer antigen 15-3 (CA 15-3), mucin-like carcinoma-associated antigen (MCA), tissue polypeptide-specific antigen (TPS), carbohydrate antigen 19-9 (CA 19-9), and prostate-specific antigen (PSA) in maternal serum (MS), umbilical cord serum (UC), and amniotic fluid (AF) and outlines their roles in the assessment of pregnancy and malignancy. All antigens studied, except CA 15-3, are oncofetal. The presence of considerable concentrations of AFP, hCG, CEA, CA125, SCC, MCA, TPS, CA 19-9, and PSA in AF during pregnancy may be attributed to their involvement in biological functions associated with fetal development, differentiation, and maturation. MS CEA, CA 15-3, and CA 19-9, in contrast to all the others, are not influenced significantly by pregnancy and thus remain reliable tumor markers in monitoring malignancy in pregnant patients.

Cytokine concentrations during the first days of life.

OBJECTIVE: To evaluate the cytokine concentration patterns during the first 5 days of life by measuring serum concentrations of type-1 cytokines, like interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) and type-2 cytokines, like IL-4, as well as the receptors of IL-2 (sIL-2R) and IL-4 (sIL-4R) during the early neonatal period. SUBJECTS AND METHODS: Forty-two healthy term neonates were included in the study. Cytokine concentrations were measured in umbilical cord, in the 1st and 5th day after birth and compared with those in serum of 30 healthy adults. RESULTS: IL-2 concentrations presented a decrease trend from umbilical cord to 5th day, while sIL-2R showed a significant elevation from umbilical cord to 5th day after birth. IL-4 concentrations did not differ significantly among umbilical cord, the 1st and the 5th day, while the sIL-4R showed the highest values in the 1st day after birth. Both IL-4 and sIL-4R concentrations in neonatal samples were elevated compared to adults. IFN-gamma concentrations increased significantly from umbilical cord to 5th day of life. CONCLUSION: Our findings indicate a dysregulation among IL-2, IL-4 and IFN-gamma concentrations during the 1st day after birth, favoring a more precocious expression of IL-2 and IL-4 against IFN-gamma that seems to be ameliorated in the end of the 1st week of life.

Vascular endothelial growth factor and placenta growth factor in intrauterine growth-restricted fetuses and neonates.

The angiogenic factors vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF) are respectively up- and downregulated by hypoxia. We aimed to study circulating levels of the above factors in intrauterine growth restriction (IUGR) and to correlate their levels with the customized centiles of the infants. The study included 25 IUGR and 25 appropriate for gestational age (AGA) full-term, singleton infants and their mothers. Maternal (MS), fetal (UC), and neonatal day 1 (N1) and 4 (N4) blood was examined. MS and N1 PlGF, as well as UC VEGF levels correlated with the customized centiles of the infants (r= 0.39, P=.007, r=0.34, P=.01, and r= -0.41, P=.004, resp). Furthermore, UC, N1, and N4 VEGF levels were higher in girls (r=0.36, P=.01, r=0.33, P=.02, and r=0.41, P=.005 resp). In conclusion, positive and negative correlations of examined factors with the customized centiles of the infant could rely on placental function and intrauterine oxygen concentrations-both being usually lower in IUGR cases-while higher VEGF levels in girls should possibly be attributed to the stimulating action of estrogens.

The influence of the mode of delivery on circulating cytokine concentrations in the perinatal period.

BACKGROUND: Cytokines play an important role during labor and full- or preterm delivery. They influence physical immunity of the fetus-neonate and express a leading role in the perinatal period, being present in maternal and fetal tissues. AIM: To investigate whether cytokine concentrations in the mother, fetus and neonate depend on the labor and the mode of the delivery. STUDY DESIGN: Prospective study. SUBJECTS: Seventy-eight healthy, non-smoking parturients (mean age 28+/-4, range 21-39 years, delivering vaginally: n=52 or by elective cesarean section: n=26) and their single, healthy, appropriate for gestational age, full-term neonates. OUTCOME MEASURES: We correlated determined circulating levels of IL-2, sIL-2R, IL-4, sIL-4R, IL-6, sIL-6R, IL-1beta, IL-8, IFN-gamma, TNF-alpha, sTNF RI, sTNF RII and RANTES in the mothers before delivery (MS), the fetuses (UC) and the neonates in days 1 (N1) and 4 (N4) of life, with the mode of delivery. RESULTS: sIL-2R in N1 and N4, sIL-4R in MS, IL-6 in MS and UC, IL-1beta in MS, UC and N1, IFN-gamma in MS and UC, TNF-alpha in UC, N1 and N4, sTNF RI in UC were significantly higher in cases of vaginal delivery than in cases of elective cesarean section (p ranging from 0.0005 to 0.05). CONCLUSIONS: Vaginal delivery promotes the production of various cytokines and their receptors, which are implicated in neonatal immunity.

Soluble Fas concentrations in the follicular fluid and oocyte-cumulus complex culture medium from women undergoing in vitro fertilization: association with oocyte maturity, fertilization, and embryo quality.

OBJECTIVE: Because soluble Fas (sFas) inhibits Fas-mediated apoptosis by preventing death signal transduction, we determined sFas concentrations in the follicular fluid (FF) and oocyte-cumulus complex culture medium (CM) from women undergoing in vitro fertilization (IVF) in order to associate its concentrations with oocyte maturity, fertilization, and embryo quality. METHODS: We studied 82 follicles from 11 healthy women (mean age, 35.4 +/- 3.8 years) using a long protocol for IVF treatment. Individual FF and matched CM samples were immediately centrifuged at 4C and sFas concentrations were determined by sandwich enzyme-linked immunosorbent assays. RESULTS: sFas concentrations were significantly higher in FF than in CM (P <.0001) and when oocytes were mature rather than immature (P <.002). Of 70 mature and 12 immature oocytes, 56 (80%) and two (16.6%), respectively, were fertilized. sFas concentrations in CM were significantly lower when mature oocytes were fertilized versus nonfertilized (P <.005). sFas concentrations in FF and CM were significantly related in an inverse manner to embryo quality (P = .004 and P = .0002, respectively). CONCLUSION: FF and CM from women undergoing IVF contain sFas. The latter has anti-apoptotic properties and levels are higher: in FF when oocytes are mature and in CM when oocytes are nonfertilized. Furthermore, FF and CM sFas concentrations are negatively correlated with embryo quality.

Age-related differentiations of Th1/Th2 cytokines in newborn infants.

OBJECTIVE: To evaluate age-related differentiation of immune response in newborns by measuring serum concentrations of interleukin-2 (IL-2), interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) during the perinatal period. SUBJECTS AND METHODS: Fifty-seven healthy term neonates, their mothers and 25 healthy adults (controls) age-matched to the mothers were included in the study. Cytokine concentrations were measured in the umbilical cord (UC), and in first-day (1N) and fifth-day (5N) neonatal samples, compared with those in maternal serum (MS) and control serum samples. RESULTS: Serum IL-2 concentrations in the UC were markedly elevated compared with those in MS and controls (p < 0.0001), decreasing significantly thereafter up to 5N (p < 0.001). IL-4 serum concentrations did not differ significantly between the UC, 1N and 5N samples; they were, however, markedly elevated compared with those in MS (p < 0.001, p < 0.0007 and p < 0.0001, respectively) and controls (p < 0.05, p < 0.01 and p < 0.006, respectively). IFN-gamma serum concentrations were significantly lower in the UC compared with those in controls (p < 0.04), increasing significantly up to 5N (p < 0.03). Both IFN-gamma/IL-2 and IFN-gamma/IL-4 ratios increased significantly in 5N, compared with those in the UC (p < 0.001 and p < 0.03). CONCLUSION: Our findings indicate a differential cytokine balance at birth with enhanced expression of IL-2 and IL-4 against IFN-gamma. However, a regularization of immune response seems to proceed quickly during the early neonatal life.

Circulating angiogenic factors during periovulation and the luteal phase of normal menstrual cycles.

OBJECTIVE: To measure serial serum concentrations of the angiogenic factors vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and angiogenin (ANG) in the periovulatory and secretory phase of normal menstrual cycles in healthy women and to determine their peaks, which might reflect the stage of their critical angiogenic action. DESIGN: Prospective study. SETTING: University departments of obstetrics and gynecology. PARTICIPANT(S): Thirty-three healthy Swedish women with regular menstrual cycles. INTERVENTION(S): Serial blood samples were collected from each woman. Luteinizing hormone surge was identified by testing morning urine. MAIN OUTCOME MEASURE(S): Circulating levels of VEGF, bFGF, and ANG. RESULT(S): Circulating peak concentrations were determined for VEGF on day 0 and 9 after ovulation, for bFGF on day 1 before ovulation and day 9 after ovulation, and for ANG on day 3 after ovulation. CONCLUSION(S): Circulating VEGF increased in a stage-dependent cyclic fashion. Basic FGF peaked during the late proliferative and mid secretory phase. Circulating ANG showed increased expression around the early secretory phase of the cycle.

Chemokines Rantes and interleukin-8 in the perinatal period: changes in serum concentrations.

Chemokines, a superfamily of polypeptide mediators, are a key component of immune surveillance and are implicated in the initiation of the inflammatory cascade. This study investigated whether serum concentrations of the chemokines regulated upon activation, normal T-cell expressed and presumably secreted (RANTES) and interleukin-8 (IL-8) change in the perinatal period because of the transition from intra- to extrauterine life, and compared determined values in mothers (MS) (n = 30) with those in their fetuses (UC), neonates (day of life 1 [N1] and 4 [N4]), and controls (CS) (n = 20). RANTES serum concentrations were higher in MS than in UC ( p < 0.006), N1 ( p < 0.0001), N4 ( p < 0.0001), and CS ( p < 0.0001). IL-8 serum concentrations in MS and UC, respectively, were significantly lower than in N1 ( p < 0.0002 and p < 0.0007) and N4 ( p < 0.0001 and p < 0.0001). Thus, after birth, neonatal serum concentrations of RANTES decrease, possibly because of elimination of the placenta (probable production site), and neonatal serum concentrations of IL-8 increase, possibly triggered by environmental antigenic stimuli to which the neonate is exposed.

In vitro fertilization: angiogenic, proliferative, and apoptotic factors in the follicular fluid.

The studies reported investigated the concentrations of angiogenic, proliferative, and apoptotic factors in the follicular fluid (FF) of individual follicles, aspirated from women undergoing controlled ovarian hyperstimulation using a long protocol for IVF treatment. Furthermore, the association of the concentrations of the preceding factors with oocyte maturity was studied. Vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), angiogenin, tissue polypeptide specific antigen (TPS), and soluble Fas (sFas) were all found in the FF of all follicles examined. Moreover, from the angiogenic factors only angiogenin concentrations, and from the apoptotic factors sFas concentrations (the soluble form expressing rather an antiapoptotic function), were positively associated with oocyte maturity, possibly indicating angiogenin's biological role beyond neovascularization and a lower apoptotic rate allowing oocytes to mature. Last, the abundant expression of TPS in FF may be indicative of intense cell proliferation, in cases of ovarian stimulation.

Soluble Fas antigen and soluble Fas ligand in early neonatal life.

BACKGROUND: After birth, apoptosis rates might slow down, compared to those in utero. Thus, factors, attenuating the apoptotic process, like the soluble forms of Fas/FasL system, may increase. AIM-STUDY DESIGN: Soluble Fas (sFas) and soluble Fas ligand (sFasL) concentrations were measured in maternal serum (MS), umbilical cord (UC) and neonatal serum in the first (1N) and fifth (5N) days after birth in order to evaluate the alterations of these molecules during the early neonatal period. SUBJECTS AND METHODS: Soluble molecules were estimated in 35 healthy, appropriate for gestational age, full-term neonates, their mothers and in 25 healthy, nonpregnant women, age-matched to the mothers (controls), using enzyme immunoassays. RESULTS: sFas concentrations in MS (p < 0.01), UC (p < 0.0001), 1N (p < 0.0003) and 5N (p < 0.02) were lower than those in controls. Neonatal sFas concentrations showed a significant increase from UC to 5N (p < 0.001). In contrast, sFasL concentrations were significantly elevated in all neonatal samples (UC, 1N and 5N) compared to those in MS and controls (p < 0.0001), showing also a significant elevation from UC to 5N (p < 0.0001). CONCLUSION: Our results demonstrate increasing serum concentrations of the soluble molecules sFas and sFasL during the first days after birth, indicating possibly a gradual decrease of apoptosis in early neonatal life.

Indicative markers of cell proliferation and apoptosis during the perinatal period.

Tissue polypeptide-specific antigen (TPS), an indicative marker of cell proliferation and soluble Fas (sFas), an antiapoptotic molecule were determined in neonatal serum (day 1-N1 and day 4-N4 of life), compared with maternal serum (MS) and umbilical cord serum (UC) to study changes of these markers during the perinatal period. Serum TPS and sFas concentrations were measured in 33 healthy, termed neonates, their mothers and 25 healthy nonpregnant controls (CS), age-matched to the mothers. TPS serum concentrations were significantly elevated in N1 and N4 as compared to CS (p < 0.0001 and p < 0.0003), increasing significantly from UC to N1 (p < 0.0001) and decreasing thereafter in N4 (p < 0.0002). MS serum concentrations, being significantly higher than CS (p < 0.0001), UC (p < 0.0001) and N4 (p < 0.003), but lower than N1 (p < 0.02) were strongly depended on the mode of delivery (p < 0.001). Serum concentrations of sFas, being lower in UC than in MS or CS (p < 0.0001), increased significantly in N4 samples (p < 0.01). A strong correlation was found between sFas serum concentrations in N1 and N4 (r = 0.65; p < 0.001). Our results demonstrate significant perinatal changes in TPS and sFas serum concentrations, possibly indicating gradual decrease of proliferation and apoptosis in early postnatal life.

Cytokine soluble receptors in perinatal and early neonatal life.

BACKGROUND: In contrast to cellular receptors, soluble receptors do not enhance the cellular activation because they do not have transmembranic and cytoplasmic parts, acting thereby as endogenous regulatory mechanisms against systemic functions of cytokines. AIM: To measure serum concentrations of the soluble interleukin-2 receptor (sIL2R), soluble interleukin-4 receptor (sIL4R), soluble interleukin-6 receptor (sIL6R), and soluble tumor necrosis factor-alpha receptor I and soluble tumor necrosis factor-alpha receptor II, during the perinatal and early neonatal period, in order to evaluate their role in activation of immune response in labor and the first days postpartum. METHODS: Soluble receptor serum concentrations were determined by enzyme-linked immunosorbent assay, in 45 healthy, full-termed neonates during the first and fifth days after birth, in 25 of their mothers (MS), in 25 samples of umbilical cords (UC) and in 25 healthy adult donors age-matched with the mothers (controls). RESULTS: Soluble receptor serum concentrations showed considerable changes during labor and early neonatal life, being significantly higher both in MS (except sIL6R) and in neonatal sample UC, first and fifth days after birth, compared with controls (p<0.0001). Neonatal serum sIL2R and sIL6R increased significantly from birth to the fifth day, while the remaining receptors showed a rapid increase in the first day (p<0.0001), declining significantly thereafter (p<0.0001). CONCLUSION: Our findings suggest that the elevated concentrations of all studied soluble cytokine receptors reflect the activation of immune response, and represent also regulatory protective mechanisms for mother and fetus-neonate against the systemic function of cytokines during labor and early neonatal life.

Concentrations of soluble Fas in maternal serum and amniotic fluid during uncomplicated pregnancies.

OBJECTIVE: To determine, during normal pregnancy, maternal serum (MS) and amniotic fluid (AF) concentrations of soluble Fas (sFas), an apoptosis-suppressing molecule that might play a role in the apoptotic process. Soluble Fas levels might explain existing immunotolerance, fetal well being, and rupture of membranes at term. METHODS: Sixty-six healthy, nonsmoking, pregnant women (mean age 32.6 +/- 4.8 years) with uncomplicated singleton pregnancies (15 in the first trimester, 30 in the second trimester, and 21 at term vaginal delivery) and 20 healthy nonpregnant women (mean age 32.5 +/- 3.8 years) were included in the study. RESULTS: MS and AF sFas concentrations were measured by a sandwich enzyme-linked immunosorbent assay, and parametric tests were used in the statistical analysis.MS and AF sFas concentrations significantly depended on gestational age (P < .0008 and P < .0002, respectively). MS concentrations were significantly lower in the first trimester than those in the second trimester (P <.003), those at term (P < .03), and those in nonpregnant controls (P < .005). AF concentrations decreased significantly at term compared with those in the second trimester (P < .0003). AF sFas concentrations in the second trimester and at term were significantly lower than respective MS concentrations (P < .00001). CONCLUSION: MS sFas concentrations decreased significantly in the first trimester of pregnancy, possibly affecting semiallograft tolerance. In the second trimester, concentrations return to control levels and remain unchanged until delivery. AF sFas concentrations decrease at term compared with the second trimester, possibly indicating increased apoptosis in preparation for rupture of membranes.

Mucin-like carcinoma-associated antigen in paired serum and breast milk samples of lactating mothers and sera of their neonates in the early postpartum period.

OBJECTIVE: To evaluate postpartum MCH changes in the early postpartum period, and to examine whether neonatal MCA is related to that in maternal serum (MS) or milk. STUDY DESIGN: MCA was measured by EIA on the second and fifth postpartum day in serum and BM from 30 lactating women and their single term neonates. Sera from 20 healthy women (controls), were also analyzed. RESULTS: All neonatal antigen concentrations were below the cut-off level for MCA (11 ng/ml). MS MCA was significantly increased compared with that in controls (P<0.00001), while antigen values in BM were highly elevated (P<0.00001), with a significant increase (P<0.0003) from the second to the fifth postpartum day. A strong correlation was found between the second and fifth day postpartum samples in MS, neonatal serum and BM MCA concentrations (r(s)=0.94, P<0.00001; r(s)=0.75, P<0.00001 and r(s)=0.69, P<0.0001, respectively). A significant correlation was also found in MCA values on the fifth postpartum day between neonatal serum and BM (r(s)=0.54, P<0.02). CONCLUSIONS: From these findings one may speculate on some ripening process in milk production and a possible transition of MCA from the neonatal gastro-intestinal tract into circulation.

Maternal serum leptin concentration during the second trimester of pregnancy: association with fetal chromosomal abnormalities.

Recent studies suggest that leptin, the product of the obese gene, is produced by the placenta during pregnancy. The present study addressed the question whether second trimester maternal serum leptin could be altered by fetal Down syndrome or Edwards syndrome. Maternal serum leptin concentrations were measured in 18 pregnancies complicated with Down syndrome, six pregnancies complicated with Edwards syndrome and 183 uncomplicated pregnancies during the second trimester of pregnancy. The present results demonstrate that leptin concentrations in uncomplicated pregnancies slightly decrease from the 16th week of pregnancy, reaching a minimum of 18.8 ng/ml around the 20th week, and then rapidly increase to 28.2 ng/ml by the 24th week. Leptin correlation with maternal body weight decreases from r=0.695 at 16-17 week of gestation to r=0.544 at >22 weeks of gestation. There was no significant difference between the mean MoMs of Down syndrome- (0.926) or Edwards syndrome- (0.960) affected pregnancies and normal pregnancies (1.002). A weak correlation (r=0.18, p<0.02) was observed between corrected leptin MoMs and human chorionic gonadotrophin (hCG) MoMs in normal pregnancies. It is assumed that around the 20th week of pregnancy placental leptin production is activated or at least is accelerated and it is added to the amount of leptin produced by maternal adipose tissue. Fetal Down syndrome or Edwards syndrome does not seem to alter maternal leptin concentration and therefore leptin cannot be used as a marker for these chromosomal abnormalities in the early second trimester of pregnancy.