Objective: Vasoconstriction during anxiety reduces fetal oxygenation and leads to hypoxia. Hypoxia in turn results in increase of the number of nucleated red blood cells (NRBCs) in the cord blood. The present study aimed to assess the effect of decreasing maternal anxiety on fetal oxygenation and NRBCs count in the cord blood.Methods:. In this study, 150 women were randomly divided into two intervention groups [supportive care and acupressure in BL32 (bladder) acupoint] and a control group . The infants' cord blood was investigated regarding the number of NRBCs and the intensity of hypoxia after birth. Then, the data were entered into the SPSS statistical software (v. 16) and analyzed using ANOVA, Chi-square test, and logistic regression analysis.Findings: The significant difference was found between the two groups regarding the number of NRBCs counted in the peripheral blood smear (P<0.001). Besides, a significant relationship was observed between the length of the first and second stages of labor and the number of NRBCs in the cord blood (P=0.01). Also, a significant association was observed between the type of delivery and the number of NRBCs in the cord blood in both intervention (P<0.001) and control groups (P=0.03).Conclusion: Doula supportive care and acupressure at BL32 point reduced the length of labor stages as well as the anxiety level. Also, nucleated red blood cells were less in the 2 groups of intervention than in control group. Regarding the fact that nucleated red blood cells cannot be the only factor for hypoxia predicting, for affirmation of this theory study with higher sample size and survey of mothers at high risk are needed. The hormones, such as catecholamines, cortisol, epinephrine, and beta-endorphins, which are secreted in response to tension and anxiety, interfere in the progress of cervical dilatation. On the other hand, they influence the uterine smooth muscles and reduce the contractile power of the uterus as well as its efficiency in the delivery process eventually lengthening the delivery, increasing the pain, and leading to anxiety[2]. During long-term anxiety, the autonomous nervous system is stimulated leading to an increase in the contraction of the smooth muscles of the arterial system. This subsequently reduces the uteroplacental blood flow as well as oxygenation to the uterus, leads to fetal hypoxia, and increases abnormal fetal heart rates[3-5]. Intrauterine hypoxia is an important risk factor of infant mortality and abnormal outcomes during infancy which causes problems in 5-10% of pregnancies. Recently, counting nucleated red blood cells (NRBCs) in every 100 white blood cells (WBCs) in infants' umbilical venous blood is considered as a sign of prenatal hypoxia. Each hypoxia event leads to a compensatory fetal reaction increasing hematopoiesis and entrance of immature red blood cells into the fetal blood flow level of which is associated with prenatal hypoxia[8]. Thus, the relationship between chronic stress and delivery outcomes, shows the necessity for intervention in order to reduce it[9]. In general, non-pharmacological methods are among the most effective methods of coping with stress and delivery pain. One of these methods involves continuous support of the mother by the doula[10]. Hung conducted a semi-experimental study on the effect of a supportive companion during labor on the delivery outcomes. The study showed that supporting the women during labor was effective in improving the uterine dysfunction and reduced the length of labor as well as the cesarean rate. Besides, the supported women felt less exhausted and were more satisfied after the delivery[11]. Teixeira et al (1991) revealed a relationship between the mother's anxiety score and increase in the vascular resistance of uterine arteries[12]. Labor anxiety and release of cortisol and catecholamines may lengthen the labor, decrease the placental blood flow, and lead to fetal hypoxia[13].Delivery is an important phenomenon and may be one of the most painful and stressful events that mothers experience during their lifetime[14]. Studies have shown that acupuncture is effective in releasing oxytocin from the pituitary gland and stimulates the secretion of oxytocin which directly stimulates the uterine contractions leading to beginning and progress of the delivery and reduction of the labor length. In general, various acupuncture points are used to induce and control the delivery and BL32 acupoint [Shang Liao (urinary bladder or bladder meridian)][17] was used in the present investigation. In this study, acupressure which is a simple, non-invasive method and doula supportive care was used in order to reduce the mothers' pain and anxiety during labor and prevent undesirable fetal outcomes, such as hypoxia. Thus, the present study aims to assess the effects of decreasing maternal anxiety on fetal oxygenation and f NRBCs count in the cord blood. Acupressure is another non-pharmacological intervention which is used in order to decrease anxiety. Acupressure is an acupuncture branch in which finger pressure is used instead of needlesThis randomized clinical trial was conducted in the delivery ward of the selected educational center of Shiraz University of Medical Sciences  in 2012. Considering d=5, α=0.05, 1-β=0.90, SD=7, and the following formula, a 150-subject sample size (50 subjects in each group) was determined for the study: Then, the subjects were selected through simple random sampling and were divided into supportive care, acupressure, and control groups using stratified block randomization. In doing so, a number was randomly selected from the table of random numbers and the researcher moved toward the right or left column or row and wrote the 5 digit numbers down. Since the participants were divided into 3 groups in this study, 3-therapy method was used and classification was performed as follows: A: supportive care group, B: acupressure group, and C: control group. Accordingly, ABC: 1, ACB: 2, BAC: 3, BCA: 4, CAB: 5, and CBA: 6. It should be noted that numbers 0, 7, and 9 were ignored. The inclusion criteria of the study were first or second pregnancy, single and term pregnancy, vertex presentation, spontaneous beginning of the delivery process, 3-4 cm cervical dilatation, being 18-35 years old, and having at least middle school degree. The exclusion criteria of the study were: suffering from any physical or mental problems, preeclampsia, having a history of smoking, maternal diabetes, Rh incompatibility, oligohyd-ramnios, and thick meconium,. [18]. In Iran, Aghamohammadi et al (2007) used this questionnaire on 150 patients undergoing surgery and reported its reliability as 97%[19] which is the basis for the present study. The supportive care and acupressure groups completed this questionnaire before and after the intervention. In the control group, on the other hand, the questionnaire was completed at the beginning of the active phase of the labor and the end of the first stage. The study data were collected using a questionnaire including demographic, clinical, and pregnancy information and an observation form including the information about the labor stages and the laboratory results. Also, Spielberger's questionnaire was employed in order to assess the anxiety level. This questionnaire includes 40 questions in two 20-item scales. The first 20 questions measure the state anxiety which is defined as a feeling of worry and tension resulting from situational stress. Yet, the second 20 questions assess the trait anxiety which refers to individual differences in the tendency toward evaluating the situations as threatening or dangerous In the first group, the researcher as the doula was beside the mother from her entering the department. She reassured the mother, encouraged her, and gave her correct and appropriate information which led to the mother's tranquility, understanding the origin of pain, developing a positive attitude toward the pain experience, and increase in her cooperation in the delivery progress.[20]. The pressure was continuously and gently applied by both thumbs for 20 minutes. It was applied by the beginning and stopped at the end of the contractions. On the other hand, the control group only received the department's routine care and underwent no interventions. In the second group, in 3-4 and 7-8cm dilatation, the mother was placed in a proper position and BL32 point was pressed. This point lays approximately one index finger length above the top of the buttock crease, approximately one thumb width either side of the spine [20]. TheP<0.05 was considered as statistically significant. After birth, the infants' cord was double-clamped before or simultaneous with the first breath. In order to facilitate the sampling, the blood was directed toward the cord by the fingers so that the umbilical cord vessels were full of blood. Then, in order to count the total number of the blood cells (CBC diff), 2cc cord blood was poured into a bottle containing Ethylen Diamine Tetracetic Acid (EDTA) and transferred to the hospital laboratory for hematological analysis. After preparing and staining the peripheral blood smear, the number of NRBCs was determined per 100 WBCs and the results were recorded in the related form. Finally, the data were entered into the SPSS statistical software (v. 16) and analyzed using Chi-square, one-way ANOVA, LSD, and logistic regression analysis. FindingsP=0.4).The study results revealed no significant difference between the study groups regarding the demographic variables including mother's age, level of education, occupation, gestational age, and number of pregnancies. In addition, the results of one-way ANOVA showed no significant difference between the three groups regarding the anxiety score before the intervention (P<0.001)  .P<0.001). In all the three study groups, 41.3% (0-10) of NRBCs of the blood cord were observed in the peripheral blood smear. Among the 9 cases with NRBCs in the supportive care group, 14% had 1 and 4% had 2 NRBCs. Also, among the 10 cases with NRBCs in the acupressure group, 10% had 3 and 2% had 1 NRBC. In the control group, on the other hand, among the 43 cases with NRBCs, 18% had 2 and 2% had 10 NRBCs. Overall, a larger number of NRBCs were detected in the peripheral blood smear in the control group compared to the two intervention groups  (OR=1.01). In all the three groups, the mean length of the first and second stages of labor was higher in the cases with NRBCs detected in their peripheral blood. In the control group, the mean length of labor in the cases with NRBCs detected in their peripheral smears was 190.0 and 175.2 minutes higher in comparison to the supportive care and acupressure groups, respectively. Among these cases, the highest and lowest mean length of labor was related to the control and supportive care groups, respectively  and the control group (P=0.03)  . MoreoveThe findings of the present study confirmed the effectiveness of applying pressure at BL32 point and psychologically supporting the mother during labor on reduction of maternal anxiety and improvement of fetal oxygenation.[21]. In the same line, the results of the study by Fassoulaki et al (2003) showed the effectiveness of acupressure in reduction of pain and stress[22]. These results were in line with those of the present study in which, the anxiety score decreased by 34.8% in the acupressure group after the intervention. Considering the supportive care group, the present study results were in agreement with those of the study by Hofmeyr et al (1991) reporting the anxiety mean score as 28.2 in the supported group and 37.8 in the routine care group[23]. In the current study also, the mean score of anxiety reduced by 37.5% in the supportive care group after the intervention which might be due to accompanying and supporting the mother leading to reduction of her pain and anxiety. Pilkington et al (2007) also expressed the effect of acupressure on the patients’ anxiety level based on Spielberger’s scale[24]. Various studies have shown that acupressure, without any complications, controls and reduces anxiety by stimulating brain responses and hormonal activities through increasing the blood flow and mediating the metabolism[25]. Therefore, it can be concluded that mother’s anxiety which is accompanied by increased uterine vessels' resistance and decreased oxygenation plays a major role in fetal as well as maternal outcomes. This emphasizes the necessity for performing interventions through labor in order to reduce the mothers’ anxiety. On the contrary, Langer et al (1998) showed that supporting the women through labor was not effective in the need for medical interventions and mothers' anxiety, pain, self-confidence, and satisfaction. In that study, the mean score of anxiety was 49.1 in the supported group and 49.2 in the control group and the difference was not statistically significant[26]. The difference between that study and the present one might be due to intervention methods, hospital policies, women's cultural background, short period of support, and types of doulas' activities. Yet the support provided by the nurses and the staff might have also been effective in reducing the difference between the two groups.After the intervention, the anxiety score decreased by 20.5 points in the acupressure group and by 20 points in the supportive care group, but increased by 11.1 points in the control group. Chao et al (2007) also performed acupressure at various points and revealed this non-pharmacological method to be effective in reduction of women's pain and anxiety during labor In this study, distribution of the NRBCs in the peripheral blood smears was 18%, 20%, and 86% in the supportive care, acupressure, and control groups, respectively. Stress and anxiety lead to secretion of epinephrine. [27]. Basically, intrauterine hypoxia is one of the main factors in increasing erythropoietin which stimulates the fetal hematopoietic system and increases the production of NRBCs. Since NRBCs can change their shape, size, and expansibility, they are released from the bone marrow into the peripheral blood. In the current study, the number of the cases with NRBCs detected in the peripheral blood smears was higher in the control group compared to the two intervention groups. Thus, fetal hypoxia was probably higher in the control group which received no interventions for reduction of pain and anxiety during labor. Several studies have indicated a significant relationship between the number of NRBCs and fetal hypoxia and have considered the number of NRBCs as an important index for identifying hypoxia[28-30]. Predicting the importance of fetal hypoxia needs awareness of its duration and grade[31]. NRBC count increases in acute and chronic hypoxia but the more the fetal hypoxia, the more increases NRBC count[29]. The average rate of nucleated red blood cells is 500 in mm3 in the first hours of healthy term infant’s life [32]. 1 to 2 percent of healthy infants have increased nucleated red blood cells without any reason[33]. For instance, in the study by Tomar et al (2011), the mean number of NRBCs was 10.34+3.87 in the fetal distress group and 5.7+2.33 in the control group[34]. In addition, Hanion-Lundberg et al (1999) measured the mean number of NRBCs as 9.2+18.1 in 1561 hypoxic infants[35]. In the study by Kovalak et al (2011) the mean number of NRBCs was 13 (range: 0-37) in the case group and 8 (range: 0-21) in the control group[36]. In the present study, providing the mothers with doula supportive care and applying pressure at BL32 acupoint were performed to decrease the effective factors in fetal hypoxia including anxiety, and prevent the undesirable fetal and maternal outcomes. However, no such interventions were carried out in the studies by Hanion-Lundberg and Kovalak. Epinephrine through beta-adrenergic receptors in the uterus leads to uterine muscle hypoxia, disruption in uteroplacental blood perfusion, and fetal hypoxia In Hanion-Lundberg’s study, NRBC count in infants with first minute’s Apgar 0-3 was significantly and statistically higher than that in infants with Apgar >7; this study included also infants with diabetic mothers and meconium stained cases. In our study, these cases and also infants with basis problem were excluded and pregnancy was terminated immediately after detection of abnormal heart models. Also, ill neonates with low Apgar that needed NICU were not in our study. Our neonates despite the abnormal heart rate model in control group had Apgar scores higher than 7 and did not need resuscitative measures and NICU care, and distress and intrauterine hypoxia intensity and duration were not high enough to cause over increase in NRBC count.+107.1) and supportive care group (227.7+33.7), respectively. As the labor length increases, mother's anxiety increases, as well. Thus, due to secretion of stress hormones, including cortisol, epinephrine, and norepinephrine, the vessels are contracted and less oxygen is delivered to the fetus. Then, the production of erythropoietin is increased in response to hypoxia and NRBCs enter the peripheral blood. A large number of studies have also shown that the infants exposed to intrauterine hypoxia have more NRBCs in their cord blood . In the studies by Ferns[39] and Lim[40] also, a significant correlation was found between the length of labor and the number of NRBCs, which is consistent with the findings of the present study. However, Kovalak[36] reported no significant relationship between the number of NRBCs and the length of labor. This difference might result from the fact that most of the control group women in Kovalak's study underwent cesarean section and, consequently, the length of labor was shorter in the control group compared to the case group (distressed infants). In contrast to the current study, Ghosh[8] did not consider hypertension, preeclampsia, tobacco use, meconium, growth restriction, and chorioamnionitis which can increase the number of NRBCs as the exclusion criteria. In this study, the labor was longer in the cases with fetal NRBCs detected in the peripheral blood smears. Besides, the highest and lowest labor length was related to the control , the mean number of NRBCs was 8.25+3.39 and 7.58+4.04 in the cesarean and natural delivery, respectively and the difference was not statistically significant[41]. Kovalak[36], Axt[33], Ghosh[8], Hanion-Lundberg[35], and Korst[28] also found no significant relationship between the number of NRBCs and type of delivery. This might be due to the fact that the above-mentioned studies had not exclude hypertension, preeclampsia, tobacco use, meconium, growth restriction, and chorioamnionitis. These factors increase the number of NRBCs regardless of the delivery mode and labor length because by uterine vessels contraction in these patients, perfusion and oxygenation to the fetus is decreased and more NRBCs are produced and entered into the peripheral blood in response to hypoxia. In this study, a larger number of NRBCs were detected in peripheral blood smears in cesarean delivery in all the three groups. This implies a significant relationship between the type of delivery and the number of NRBCs in the blood cord in both the intervention groups and the control group. In line with these results, FernsThe findings of the present study showed that supportive care and acupressure were effective, safe, simple, and inexpensive techniques which could be used in order to reduce the delivery pain, mother’s anxiety, and length of labor, resulting in improvement of oxygenation to the fetus and prevention of asphyxia. Asphyxia, regardless of duration, does not always increase the NRBC count and NRBC count is far more increased in some cases without asphyxia. Therefore, NRBC count is not the only decisive factor of intrauterine asphyxia intensity and duration. In case a large number of NRBCs are detected in infants, they should be provided with more care in order to prevent them from probable nervous damages. Although the number of NRBCs alone cannot be a prognostic factor, further studies on this issue can confirm or reject the results obtained in the current study. Overall, considering the national and international approach toward physiological delivery, this non-pharmacological method is recommended to be used to reduce the mothers' anxiety during labor and improve the maternal and fetal outcomes. One of the limitations of this study was interference of gynecologists and midwives in the normal process of labor. In case their interventions affected the results, the patient was excluded from the study. Moreover, the participants’ mental conditions while completing the questionnaire, family problems, and undiagnosed diseases could affect the study results, which was out of the researcher’s control.Z. Masoudi, M Akbarzadeh, F. Vaziri and M. Ramzi conceived of the study and participated in its design and coordination and helped to draft the manuscript. Z. Masoudi collected the data. Z. Masoudi and N. Zare performed the statistical analysis. Z. Masoudi and M. Akbarzadeh made critical revisions to the paper and translated it into English language. All authors read and approved the final version of the paper.


The aim of the present study was to evaluate the clinical value of magnetic resonance imaging (MRI) in the preoperative T staging of gastric cancer and in the postoperative pathological diagnosis. In total, 30 patients with gastric cancer were investigated, including 19 males and 11 females . The preoperative depth of invasion (T stage) was evaluated according to the characteristics of the imaging performance. The evaluation results for the MRI T staging were as follows: T1 stage accuracy, 90% with a specificity of 96% and sensitivity of 60% ; T2 stage accuracy, 86.7% with a specificity of 87.5% and sensitivity of 83.3% ; T3 stage accuracy, 90% with a specificity and sensitivity of 90% ; and T4 stage accuracy, 96.7% with a specificity of 100% and sensitivity of 87.5% . The results demonstrated that, with reference to pathological diagnosis, the MRI method exhibited high accuracy, specificity and sensitivity in determining the preoperative T stage in gastric cancer patients. Gastric cancer is one of the most common types of gastrointestinal cancer in China, with notably high incidence (3.3%) and mortality (75%) rates, and is associated with a poor prognosis. Therefore, the accurate preoperative evaluation of the extent of tumor tissue infiltration may be extremely valuable.Previously, the diagnosis of gastric cancer has predominantly been based on upper gastroenterography and gastroscopy, which directly or indirectly observes the morphology, range and pathological changes occurring on the gastric mucosal surface. These were the necessary methods to confirm the identification, location and characteristic diagnosis of gastric cancer. These methods were unable to directly reveal the stomach structure, were limited in identifying the invasion depth of the gastric wall, extra-stomach infiltration and metastasis, however, they aided to a certain extent with the qualitative diagnosis of cavity lesions . Recentlet al , the accuracy of preoperative TNM Classification of Malignant Tumours (TNM) staging in gastric cancer has gradually improved and exhibited its superiority . Matsushet al  previousl  were included. Prior to scanning, endoscopy was performed as the diagnostic test. The 30 patients included 19 males and 11 females  and there were 12 cases of gastric cardia tumor, 10 cases of gastric body tumor, four cases of gastric antrum tumor, and four cases of gastric antrum and body tumors. All of the cases were confirmed by pathology and included 15 cases of poorly differentiated adenocarcinoma, 12 cases of moderately and well-differentiated adenocarcinoma, one case of undifferentiated adenocarcinoma, one case of signet ring cell carcinoma and one case of mucinous adenocarcinoma. The 30 patients received a preoperative MRI examination one week prior to surgery and the results were assessed and identified by two experienced radiologists. The current study was conducted in accordance with the declaration of Helsinki and approved by the Ethics Committee of the Handan Hospital of Jizhong Energy Fengfeng Group . Written informed consent was obtained from all patients.The Siemens 1.5T Tim Avanto MRI instrument  was used. Anisodamine  was intramuscularly injected and 500 ml warm water was administered orally 15–30 min prior to the examination. An additional 500 ml warm water was administered prior to the patient lying on the examination table; the patients were generally in the supine position to make a major cross section. Conventional scanning with T1-weighted imaging (WI), T2WI, short inversion time inversion recovery, diffusion weighted imaging and enhanced scanning were performed. A gadopentetate dimeglumine injection  was used for the enhanced scanning and the scan range was between the top of the diaphragm and the umbilicus. Three-phase dynamic enhancement was performed in the arterial phase (25–30 sec), the venous phase (65–70 sec) and the equilibrium phase (3–4 min), following initiation of the injection. A scan voltage of 120 kV and a current of 120 mA were used.th edition) TNM staging of gastric cancer was adopted: T1 stage, no thickening of the gastric wall, with abnormal enhancement of the stomach lining and enhanced tissues not exceeding the intermediate layer; T2 stage, abnormal thickening of the stomach wall, while the whole outer layer of the multilayer-structure of the stomach wall is structurally integrated or the serosal surface is smooth and tidy; T3 stage, the whole stomach is infiltrated by the tumor and the external edge of the stomach wall or peripheral-stomach adipose tissue exhibits lower or irregular signal patterns or interruption; and T4 stage, structure signals of the stomach-adjacent organs are changed or an abnormal enhancement shadow of the stomach-adjacent organs appears in the enhanced scan.The following 2009 Union for International Cancer Control .In total, four cases of T1 stage , eight cIn total, five cases of T1, six cases of T2, 10 cases of T3 and eight cases of T4 stage were identified by postoperative pathology.MRI correctly diagnosed three cases, misdiagnosed one case and missed two cases. The MRI diagnostic accuracy of T1 stage was 90%, with a specificity of 96% and sensitivity of 60%  and missed one case. The underestimated case was diagnosed as T3 stage due to the indistinct appearance of the external edge, while the two overestimated cases were diagnosed as T1 stage, as the thin wall induced stronger signals. The MRI diagnostic accuracy of T2 stage was 86.7%, with a specificity of 87.5% and sensitivity of 83.3%  and missed one case. The underestimated case was diagnosed as T4 stage due to the ambiguous expansion of the fat surrounding the lesion, while the overestimated case was diagnosed as T2 stage due to the integrated and continuous edge of the stomach wall. The MRI diagnostic accuracy, specificity and sensitivity of T3 stage were all 90%  compared with the postoperative pathological observations, particularly for the diagnosis of T3 and T4 stages. The κ values were >0.75 for the T3 and T4 stages, which reflected a relatively satisfactory degree of consistency between the two diagnostic methods.Commonly, gastric cancer occurs in the gastric antrum, predominantly in the lesser curvature of the stomach, which accounts for ~75% of gastric cancers. Other types are located in the fundus (including the cardia) or gastric body and, according to the development of the gastric cancer, it may be categorized as early- or advanced-type. Early gastric cancer is defined by tumor tissues infiltrating only the lamina propria and submucosa, not the stomach muscle layer. Minute gastric carcinoma is an early gastric cancer, with a diameter of <5 mm; small gastric cancers exhibit diameters of 6–10 mm. The five-year survival rate of early gastric cancer is ≥85% and that of minute gastric cancer may be ~100%, therefore, the early diagnosis and treatment of these types of cancer is important . Next, the contrast agent gradually flows out of the vessel into the extravascular space  and finally achieves the equilibrium phase. The gastric blood supply is rich, with the main blood supply from the left gastric artery, hepatic artery and pulmonary artery, arising from the celiac trunk. The left gastric artery supplies the lesser curvature of the stomach, the left and right gastroepiploic arteries supply the greater curvature of the stomach and the short gastric arteries supply the fundus of the stomach. Angiography of gastric cancer and microvascular studies have shown that neovascularization may be observed in the arterial and capillary phases of the majority of gastric cancer, with a rich blood supply, a high number of distorted new blood vessels and an increased vascular volume . Additioet al  exhibited a high reinforced magnitude of the mucosal layer with a linear shape in the arterial phase. However, this was more evident in the parenchymal phase, in which the submucosa was continuous and complete. By contrast, the significant enhancement effect disappeared in the equilibrium phase. In the 26 advanced gastric cancer cases, a marked thickening of the inner layer was observed in the arterial phase and a gradually expanding enhancement area appeared throughout the whole lesion in the parenchymal phase.et al  A long clinical assessment time, normally ~30–45 min; ii) a small and limited scan range; and iii) poor image quality, as dynamic enhanced scanning requires the patients to breathe repeatedly, which certain patients are unable to cope with, thus producing an unclear image. However, the continued development of MRI technology may resolve these difficulties and increase the value of adopting MRI for the preoperative staging of gastric cancer.


A functional national health research system (NHRS) is crucial in strengthening a country’s health system to promote, restore and maintain the health status of its population. Progress towards the goal of universal health coverage in the post-2015 sustainable development agenda will be difficult for African countries without strengthening of their NHRS to yield the required evidence for decision-making. This study aims to develop a barometer to facilitate monitoring of the development and performance of NHRSs in the African Region of WHO.The African national health research systems barometer algorithm was developed in response to a recommendation of the African Advisory Committee for Health Research and Development of WHO. Survey data collected from all the 47 Member States in the WHO African Region using a questionnaire were entered into an Excel spreadsheet and analysed. The barometer scores for each country were calculated and the performance interpreted according to a set of values ranging from 0% to 100%.The overall NHRS barometer score for the African Region was 42%, which is below the average of 50%. Among the 47 countries, the average NHRS performance was less than 20% in 10 countries, 20–40% in 11 countries, 41–60% in 16 countries, 61–80% in nine countries, and over 80% in one country. The performance of NHRSs in 30 (64%) countries was below 50%.An African NHRS barometer with four functions and 17 sub-functions was developed to identify the gaps in and facilitate monitoring of NHRS development and performance. The NHRS scores for the individual sub-functions can guide policymakers to locate sources of poor performance and to design interventions to address them.The online version of this article (doi:10.1186/s12961-016-0121-4) contains supplementary material, which is available to authorized users. The African Region of WHO bears a double burden of communicable and non-communicable diseases, and many of its countries have not achieved the health Millennium Development Goals (MDGs) . Health An NHRS is defined as the people, institutions and activities whose primary purpose is to generate and promote the utilisation of high quality scientific knowledge to promote, restore and/or maintain the health of a population . Its goaGovernance is an overarching function of the government to ensure effective oversight, coalition building, system design, accountability, and regulation of all R4H taking place in both the public and private sectors. The indicators for R4H governance include availability and implementation of a national R4H policy and strategic plan, a R4H agenda, a R4H legislation or law , a R4H focal point, and a functional ethical review committee to protect the dignity, integrity and safety of research participants.The developing and sustaining R4H resources function covers building or strengthening and sustaining of human resources for biomedical, bioscience, epidemiology, public health, social science and health system research, physical infrastructure, and institutional and systemic capacities to manage knowledge . In thisThe function of producing and using R4H entails the conduct, dissemination and application of R4H in advocacy, policy and planning, systems strengthening, programme implementation, development of new products and tools, monitoring and evaluation, etc. , 4. The The existence of a knowledge platform that is designed for translating, synthesising and communicating research to inform health policy and practice could be used as a proxy for the use of research findings. In this paper, the parameters for the production and use of the research function include the existence of a R4H programme action plan, a knowledge translation platform and a national R4H management forum, and the number of R4H articles published per person.Financing of R4H refers to the estimation of recurrent and capital cost of R4H; mobilisation/collection of R4H funds from individuals, businesses, the government and donors; accumulation and management of R4H funds; allocation of funds to individuals, institutions and networks within NHRSs that govern R4H, create R4H inputs, produce R4H, and monitor and evaluate R4H; and tracking of expenditures on R4H. The parameters for the financing function include the ministry of health’s budget line for R4H and the government’s allocation of 2% of the national health budget to R4H, per the Algiers Declaration of Health Ministers in 2008 .As the world moves towards the post-2015 sustainable development agenda , which iThe 28th session of the WHO African Advisory Committee for Health Research and Development in 2014 recommended the development of a barometer to help in assessing and tracking of the performance of NHRSs in countries of the WHO African Region . This paThe proposed barometer will have four functions, derived from the standard NHRS conceptual framework . These aIn this paper, governance has six sub-functions, whereas developing and sustaining resources has five, producing and using research has four, and financing has two, making a total of 17 sub-functions  component had five instead of 15 questions – those enquiring when it was started, under which ministry, information and communication technologies connectivity, and regarding WHO Collaborating Centres were omitted. Health Research Financing and Budget had four instead of seven questions – queries on total government budget and total spending on research in general from all sources were missing. The rationale for using the short version was to reduce the amount of time needed for completion and to increase the response rate. The majority of questions omitted were those that entailed qualitative subjective judgements of respondents. In the NHRS assessments conducted in the region in 2003 and 2009 using the long questionnaire, only 10 and 44 countries, respectively, responded. We believe that the use of the short version of the questionnaire contributed to having completed questionnaires from all the 47 Member States in the WHO African region. We have no reason to believe that the omitted questions influenced the values of the indices for sub-functions and the overall composite index. Since the questions in the 2014 short version questionnaire are those used in the previous two surveys, authors did not think there was need for pilot testing or pretesting.Health Research Policy and Systems [The questionnaire was sent by email to all the 47 Member States in the WHO African Region through the WHO country representatives. It was accompanied with a covering memo from the WHO Regional Director underscoring the urgency of the survey and asking the country representatives to personally facilitate and follow-up on the survey. The national health research focal person in each of the countries, irrespective of the government ministry hosting them, had the primary responsibility for completing the questionnaire. The WHO country office national professional officer in charge of research hand-delivered the questionnaire to the national focal point, provided a briefing on the survey, went through the questionnaire with a national focal point and closely monitored the progress on the completion of the questionnaire. All the 47 countries completed and returned the questionnaire to the Regional Office for Africa. Excerpts of the questions related to the sub-functions are contained in Box  Systems .Box 2QueAll the data used in this study were collected using the questionnaire mentioned above. The questionnaire did not, however, capture data on the total peer-reviewed R4H articles per country in the Region, as most of the countries did not have a repository with that information. Therefore, those data were obtained from a bibliometric study published by some of the authors of this article .Taking into account the information from the completed questionnaires on each country’s NHRS (step 3), we assessed the dichotomous/categorical sub-functions and allocated them a percentage score ranging from 0% for non-existence to 100% for existence. For example, if with regards to national policy on research for health, a respondent answered ‘yes’ to the question ‘Does the country have a valid official health research policy?’, that country received a score of 100% for that sub-function, while a country that had ‘no’ for an answer received 0%. This exercise was undertaken for all the questions with a ‘yes’ or a ‘no’ answer.For sub-functions with continuous value answers, the score was the actual value. For example, for the number of technical and support staff sub-function question ‘How many technical and support staff are there in the programme?’ the actual value was the number of staff per 100,000 population, that is staff density. This value was obtained by dividing the total number of staff by the population and multiplying the outcome by 100,000. Similarly, the actual value for R4H peer-reviewed publications per 100,000 population was estimated through the division of the total number of R4H articles published by the population and multiplying the outcome by 100,000.Another example of a sub-function with a continuous value is the number of universities with faculties of health sciences (or medicine), where the actual value was the number of universities with faculties of health sciences per 1 million population. This value was calculated by dividing the number of national universities with faculties of health sciences or medicine by the country’s population and multiplying the outcome by 1 million.Concerning the sub-function entitled ‘progress towards the target of allocating 2% of the national health budget on R4H’, the score was the ratio of the money budgeted by the Ministry of Health (MOH) for research in the last financial year (2012/2013) to the overall MOH budget for that year multiplied by 100%.i is the ith sub-function, such as the existence of a national policy on research for health (R4HP), a strategic plan on research for health, a national research for health priority list/agenda, a national ethics review committee, etc. The formulae used for calculating the indices for the sub-functions are very similar to those used by the United Nations Development Programme to calculate the human development index [Actual R4HP is the actual research for health policy score, Minimum R4HP is the minimum research for health policy score, and Maximum R4HP is the maximum research for health policy score. As an example, if we assume that the regional minimum R4HP score is 0 and the maximum score is 100, a country like Uganda would have an actual score of 100, since it has an R4HP. The R4HPI for Uganda would be obtained as follows:All the indices for the 17 individual sub-functions were calculated using the following general formula:nt index  and the nt index . For insActual NHB is the budget allocated for R4H score  to the overall MOH budget multiplied by 100%), Minimum NHB is the minimum budget allocated for R4H score in the Region, and Maximum NHB is the maximum budget allocated for R4H score in the Region. For example, if we assume that the regional minimum NHB score is 0.0104 , the maximum NHB score will be 3.2403 (Tanzania’s score), and the actual NHB score for Uganda will be 0.46% . The NHBI for Uganda can be obtained as follows:Similarly, the index for progress towards the target of allocating 2% of the national health budget (NHB) to R4H (NHBI) was estimated as follows:Actual PPC is the R4H publications per 100,000 population score , Minimum PPC is the minimum number of publications per 100,000 population in the Region, and Maximum PPC is the maximum publications per 100,000 population in the Region. For example, if we assume that the regional minimum PPC score to be 0.026225 (Democratic Republic of the Congo’s score) and the maximum PPC score to be 14.9 (Seychelles’ score), and we consider that the actual PPC score for South Africa is 5.9806, the PPCI for South Africa can be obtained as follows:The index for R4H peer-reviewed publications per 100,000 population (PPCI) was estimated as follows:The indices for all the sub-functions were estimated in a similar way.NHRSBScore) for each country as follows:TNSF is the total R4H sub-functions, which is 17 in this study. The national NHRSBScore is measured on a scale of 0 (or 0%) to 1 (or 100%). A barometer score of 0% denotes that NHRS does not exist, 1% to 49% indicates that NHRS performance is below average, 50% suggests that NHRS performance is average, 51% to 99% shows that NHRS performance is above average, and 100% implies that NHRS is performing optimally.After appraising the individual sub-function indices, we calculated the overall NHRS barometer score  was calculated using the following formula:RHRPIAVER Score is the average regional health research policy index, NHRPI is the national health research policy index, N is the total number of Member States in the African Region, which is 47.The average regional health research policy index  to 1 (or 100%), a barometer score of 0.49 (or 49%), for example, indicates that more than half of the Member States did not have a national health research policy. The regional indices for the other sub-functions were calculated in the same manner.Since the RHRSBScore) or index is equal to the sum of the average regional indices for the 17 sub-functions divided by 17. This can be expressed symbolically as follows:TNSF is the total health research system sub-functions included in the barometer, which are regional health research policy index (RHRPI), regional health research law index (RHRLI), regional strategic health research plan index (RSHRPI), regional health research management forum index (RHRMFI), regional ethical review committee index (RERCI), regional health research priority list index (RHRPLI), regional health research focal point index (RHRFPI), regional universities with faculties of health sciences/medicine index (RUFHSI), regional health research institutes or council index (RHRCI), regional R4H programme index (RHRPRI), regional R4H programme staff index (RHRHRI), regional R4H programme action plan index (RHRPAI), regional knowledge translation platform index (RKTPI), regional budget line for R4H index (RBLHRI), regional government spending on R4H index (RHRBI), regional non-governmental organisations R4H index (RNGOI), and regional publications per 100,000 population index (RPPCI).The overall regional health research system barometer score  hand-delivered the questionnaire and explained the aims of the survey; (2) gave assurance of the confidentiality of the process and that the survey did not entail any risks; (3) clarified that no benefits would accrue to respondents individually but the survey would inform the development of a NHRS barometer and a regional strategy; and (4) specified the right to refuse to participate in the survey and that refusal to do so would not have a negative impact of any kind. The informed consent statement was included in the preamble to the survey questionnaire, and the respondents were required to sign it.RHRSBScore), the average score for each of the research system functions, and the average score for each of the sub-functions. The overall R4H barometer score for the Region was 0.42 , which indicates that the performance of NHRSs in the African Region is below average. To understand the source of the poor score, the performance of each NHRS function and its related sub-functions was further examined.The Region’s R4H systems were categorised as below average if their barometer score was less than 50%, average if the score was 50%, and above average if the score was over 50%. Table The average regional score for governance of R4H was 61%, which is above average. Within governance, the indices for ethical review committees and the health research focal points were both above 80%, but those for health research policy, research law, and strategic health research plan were all below average.The mean regional score for this function was 35%, which is far below average. The indices for health research institutes (or council) and R4H programme were slightly above 50%, but those for universities with faculties of health sciences and R4H programme staff density were below average, with the latter being extremely low at 0.06%. This depicts the acute shortage of human resources for health research in the African Region.The average score for producing and using research was 32%. The index for health research management forum was above average, but those for R4H programme action plan, knowledge translation platform and publications per 100,000 population were less than 35%.The financing of R4H function had a mean score of 27%. The index for the budget line for R4H was 47% and that for government allocating 2% of national health budget to R4H was only 6%.Among the 47 countries, the average performance of NHRS was less than 20% for 10 countries, 20–40% for 11, 41–60% for 16, 61–80% for nine, and over 80% for one. The performance of NHRS in 30 (64%) of the countries in the Region was below average. Additional file To assess NHRS performance based on income, the countries were categorised under three income groups: Group 1 consisted of nine high- and upper middle-income countries, group 2 of 13 lower middle-income countries, and group 3 of 25 low-income countries (Table To determine the relationship between the NHRSB score and the gross domestic product (GDP) per capita, a regression was undertaken of the logarithm of NHRSB scores (logTHEPC) against the logarithm of GDP per capita (logGDPPC) to test the null hypothesis that there was no statistically significant relationship at 95% confidence level. The coefficient of determination (R-squared) was 0.0559, implying that GDP per capita explained only about 6% of the total variation in the NHRSB scores. The results of the log-linear regression were as follows:logGDPPC was less than the critical ‘t’ value of 1.960 (from the Student’s t-distribution table) at 95% confidence level (a significance level of 5%), we accept the null hypothesis that there is no statistically significant relationship between the GDP per capita and the NHRSB score. The coefficient for GDP per capita was −0.133, implying that, on average, a unit percentage increase in the GDP per capita will result in a −0.133 percentage decline in NHRSB. This means that, in African countries, GDP per capita currently does not influence NHRS performance.Since the computed ‘t’ value (1.63) of This paper has developed an African health systems barometer as a tool for use in identifying gaps in monitoring of NHRS development and performance in the Region. The barometer consists of the four standard functions of an NHRS and 17 sub-functions. The country and regional sub-function NHRS barometer scores would be useful in monitoring the progress in the implementation of the new African Region’s strategy on research for health that was adopted in November 2015 by the ministers of health .As reported by Kirigia et al. , betweenThe overall average NHRS performance in the African Region was 42% (standard deviation = 20) as was the median score, which was below average. The NHRS score varied widely, ranging from 6% for Sao Tome and Principe, a lower middle-income country, to 81% for Rwanda, a low-income country. In the NHRSB score ranking based on 2014 data, the top five countries were Rwanda, Senegal, South Africa, Tanzania and Uganda, each with a score above 70%. The bottom five countries were Equatorial Guinea, Chad, Seychelles, South Sudan, and Sao Tome and Principe, each with a score below 18%.Our finding that GDP does not significantly or positively influence NHRSB performance is of interest and with implications. One implication is that there is poor allocation of existing funds to R4H, which might be due to factors such as lack of political will, inexistence of R4H governance structures , and unawareness of the positive link between R4H and health development with economic prosperity . Thus, aA few limitations of this study warrant clarification. The NHRSB suffers the same kind of limitations inherent in other types of indices such as the human development index , 30. It The sub-function on R4H publications per 100,000 population took into account only articles indexed in PubMed and other international databases . It exclFinally, the questionnaire used to collect the data on most of the sub-functions had dichotomous (yes or no) questions, which would not capture the qualities of the variables assessed. For example, it is likely that the quality of research policies or any other sub-function of NHRS will vary among the countries. Further, the data on spending on R4H were not completed in the questionnaire for many countries.Though NHRSB is far from perfect, it provides an indication of the performance of an NHRS in terms of sub-function scores. An objective parameter to identify gaps and monitor progress is always useful, particularly when the same tools are used to compare different time points in the same region or country. It is widely believed that, apart from the enormous investments by countries and international partners, periodic assessment of country performance on the MDGs helped to identify the parameters that needed strengthening and induced positive competition amongst countries. Given the importance of health research, particularly in the African Region, to optimise the use of existing interventions, identify new tools and assess progress towards the newly embarked-on goal of universal health coverage, any element that can enhance R4H would be useful.In spite of the abovementioned limitations, the NHRSB has a number of advantages: (1) it combines estimates for all the 17 sub-functions of NHRS, spanning the four functions of NHRS and, thus, generating a wide range of information; (2) it uses data that are likely to be readily available in the public domain such as government reports and offices; (3) it allows countries to be ranked in order of their achievements in NHRS development; (4) it allows the comparison of the performance of one sub-function against another; (5) it can be used to help diagnose the sources of weaknesses within an NHRS; (6) it provides a framework for scoring the performance of functions and sub-functions of NHRS; (7) it is largely factual since it does not weigh sub-function scores, which would engender subjectivity; (8) it can be used for cross-country comparisons and benchmarking and for tracking individual countries’ progress over time; and (9) it can highlight the countries with different NHRSB scores but the same per capita GDP, which may imply lurking inefficiency.The African region strategy on research for health recommends that countries establish national health research and development observatories and registries to facilitate setting of health research priorities . For exaWe suggest four areas for future research. First, criteria for assessing the quality of specific sub-functions in individual countries should be developed and key informants should use the criteria to score the sub-functions on a scale of 0% to 100%. Second, a survey to determine whether the sub-functions of the NHRS barometer should be weighted should be conducted. Third, R4H accounts should be performed to obtain comprehensive estimates of domestic and external resources spent on NHRS using, for example, the methodology proposed in the Organization of Economic Cooperation and Development Frascati manual , 33. FouEvidence is urgently needed to guide African countries in their quest for universal health coverage in the context of the unfinished health MDG agenda and the post-2015 Sustainable Development Goals. Generation of appropriate home-grown evidence calls for the existence of effectively performing NHRSs. The weakest NHRS indices in the Region are R4H human resources, government spending on R4H, publications in peer-reviewed journals, and availability of institutions conducting health research. NHRS scores for individual sub-functions can guide policymakers to locate the sources of poor performance so that interventions for their improvement can be designed. The scores will also facilitate the monitoring of NHRS development and performance in the Region.


Optical fiber sensors, thanks to their compactness, fast response and real-time measurements, have a large impact in the fields of life science research, drug discovery and medical diagnostics. In recent years, advances in nanotechnology have resulted in the development of nanotools, capable of entering the single cell, resulting in new nanobiosensors useful for the detection of biomolecules inside living cells. In this paper, we provide an application of a nanotip coupled with molecular beacons (MBs) for the detection of DNA. The MBs were characterized by hybridization studies with a complementary target to prove their functionality both free in solution and immobilized onto a solid support. The solid support chosen as substrate for the immobilization of the MBs was a 30 nm tapered tip of an optical fiber, fabricated by chemical etching. With this set-up promising results were obtained and a limit of detection (LOD) of 0.57 nM was reached, opening up the possibility of using the proposed nanotip to detect mRNAs inside the cytoplasm of living cells. Usually, information on basic cellular signaling processes associated with diseases is not directly deducible from measurements on an average of a pool of cells. In fact, new information can only be obtained by monitoring the cellular signaling pathways directly inside intact cells . TraditiAmong these nanotools, nanoprobes based on fiber-optical nanotips were conceived as integrated nanodevices consisting of a recognition molecule coupled to the optical transducing element  interfaced to a photo- or spectrometric detection system, aimed to perform diagnosis within single living cells ,13,14,15et al. [Chemical nanosensors were first developed for monitoring calcium and nitric oxide, and other physiochemical parameters in single cells , showinget al.  preparedet al.  developeet al. ,20,21,22et al.  reportedIn the field of DNA/RNA analysis in cell, MBs were proposed by Tyagi and Kramer ,27,28,29A great many fiber nanotips are characterized by a metal coating which allows them to illuminate exactly only the fiber nanotip. Actually, it is very difficult to quantify the exact fraction of light that will reach the nanotip since, below a certain inner diameter, even the lowest guided mode runs into cutoff, where the wave vector becomes imaginary and thus the mode field decays exponentially . HoweverIn this paper we present preliminary results concerning the use of an optical fiber nanotip coupled to a MB for DNA detection . The solIn particular, the attention was focused on the mRNA for survivin, a protein highly expressed in most types of cancer. In this case, the MB could act not only as detector of the over-expression of the mRNA for the survivin (diagnosis) but also as the blocking agent of the synthesis of the protein itself (therapy), as we demonstrated with Real-time PCR and western blotting experiments which showed a time-dependent reduction of survivin mRNA and protein after 100 nM-MB treatment, respectively, with the molecular beacon transfected into A375 melanoma cells ,32.After the characterization of the sole MB in solution carried out to verify the reliability and the effectiveness of the fluorophore/quencher pair immobilized at the extremities of the single-stranded DNA and at the same time of the good interaction of the MB with its target, a thorough opto-chemical characterization was performed in order to evaluate the possibility of getting a reliable, reproducible and robust system consisting of the MB immobilized onto the fiber nanotip.2) for the preparation of tris-buffer ; hydrofluoric acid (HF); hydrochloric acid (HCl); sulfuric acid (H2SO4); hydrogen peroxide (H2O2); methanol (MeOH) were purchased from Sigma-Aldrich . (3-Aminopropyl)triethoxysilane (APTES) was purchased by abcr GmbH . N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) was purchased from Pierce . The molecular beacon for survivin, its complementary target sequence, the non-specific random sequence and the labeled linear probe were purchased from IBA .All the reagents for the preparation of phosphate buffered saline (PBS), 40 mM, pH 7.4; tris hydrochloride (tris-HCl), magnesium chloride  and BlackBerry® Quencher 650 , was chosen with respect to the one used for the published MBs in order to obtain a greater quantum yield (QY = 65 of the ATTO647N instead of 0.28 of the Cy5). The sequences of the different oligonucleotides  are reported below:MB     5'-(ATTO647N)CGACGGAGAAAGGGCTGCCACGXCG(BBQ)-3' X=C6-dT ThioTarget    5'-CCCCTGCCTGGCAGCCCTTTCTCAAGGACC-3'Random sequence 5'-ATCGGTGCGCTTGTCG-3'Linear probe  5'-(ATTO647N)GAGAAAGGGCTGCCA(Thiol)-3'The sequence for the survivin molecular beacon  was chosAll the measurement carried out with both the MB in solution and the MB immobilized on the fiber tip were carried out at room temperature.The optical fiber nanotip was manufactured by a dynamic chemical etching method ,38, by mUsing different dynamic regimes of the mechanical movements during the chemical etching process, it was possible to vary the cone angle, the shape and the roughness of the nanoprobes. It was found that the tip profiles were determined by the nonlinear dynamic evolution of the meniscus of the etching solution near the fiber. Different regimes can be generated by changing the ratio between the angular velocities and the ratio between the radii of optical fiber and vial, ranging from laminar flow to the onset of chaotic flow. The type of flow of the viscous HF solution caused by the rotation of the vial and the optical fiber can be described in the framework of the Taylor-Couette flow (TCF) theory ,40.Moreover, by an accurate control of the extraction speed of the fiber from the HF solution, the length and the angle of the nanotip were precisely controlled, and the capillarity effects at the nanoscale were minimized by limiting strong friction on the optical fiber nanotip in the final part of the etching process.SEM pictures and AFM measurements have been performed on different nanotips showing our ability to reproduce typical tip features, such as short taper length (~200 μm), large cone angle (from 15° to 40°), small probe tip dimension (less than 30 nm), and roughness below 10 nm. The geometrical characterization demonstrated that, with this method, a high yield of reproducible nanotips can be obtained, overcoming some drawbacks of conventional etching techniques. The nanometric roughness is the key point to keep the scattering, which can perturb the measurements, at the lowest levels. In particular, in the left top part a general view of the short taper length and the large cone angle (about 40° in this example) of the nanotip is shown, while in the left bottom an AFM image (10 µm × 10 µm) is reported, with an estimated roughness of the tip of about 10 nm. On the right panel is reported a zoom on the distal end, showing the apex angle (about 24°).2SO4:H2O2 7:3) for 30 min to carry out a strong cleaning and to activate the silanol groups present on the surface. Then the tapered fiber nanotip was functionalized by a 2-hour long silanization procedure using 5% APTES in MeOH/H2O (1:1) in order to achieve the amino groups onto the surface of the nanotip. After several rinsing cycles in MeOH and deionized water, the nanotip was cured in oven at 100 °C for 1 h.The glass nanotip was dipped in piranha solution  and then left to slowly equilibrate down to room temperature in order to optimize the hairpin formation. The immobilization of 1 µM MB (in tris-buffer), or of 1 µM linear probe, onto the nanotip was performed over night at 4 °C.et al. [Scanning Electron Microscope  was used to characterize the geometrical features of the nanotips, whereas the surface roughness was checked by Atomic Force Microscopy .An in-house optical set-up, illustrated in The experimental set-up for fluorescence measurement of the molecular beacon in solution was very similar to the one described in The functionality of the MB was firstly investigated in buffer solution by hybridization with the specific target at different incubation times. Different buffers and different culture media were already tested and reported, such as Dulbecco’s modified Eagle’s Medium (DMEM) ,44. Evenex 635 nm) of the MB 100 nM in Tris buffer alone and after 1, 3 and 5 h of incubation with the specific target 0.2 µM . Before working with the molecular beacon, the fluorescent linear probe was immobilized onto the nanotip in order to evaluate the actual possibility of sensing the presence of a labeled oligonucleotide by direct excitation of the oligonucleotide fluorophore by means of the same fiber nanotip. After the probe immobilization, the fluorescence of the ATTO647N fluorophore labeling the probe, was measured  with a standard deviation of 436 (a.u.) leading to a signal variability of 3.8%.In The performance of the fiber nanotip to act as sensor was evaluated by monitoring the fluorescence signal as a function of time, when exposed to different concentrations of the target. Its specificity when interacting with the random sequence was also investigated . In partThe curve in The limit of detection achieved opens up the possibility of using the proposed nanotip to detect mRNAs inside the cytoplasm of living cells. In fact, the concentration of specific mRNAs has been estimated to be in the nM range , althougBy investigating the possibility of regenerating the nanotip for multiple use by the use of HCl, we discovered that it is possible to increase the performance of the sensor. An 8-fold increase in the fluorescence signal due to the interaction of the MB with the target can be observed. This behaviour can be ascribed to the unfolding effect of HCl which is able to open the hairpin structure of the MB and the subsequent refolding of the beacon hairpin structure in a more efficient manner with respect to the untreated beacon which could be also folded around the nanotip and consequently hindered towards the hybridisation with the target. This fact can provide the explanation of the faster kinetics which is observed after the HCl treatment, with the plateau reached after only 4 min, instead of 15 min. The use of HCl 1–2 mM, among others regenerating solutions, is quite common in DNA biosensor regeneration ,46,47. TThe present paper describes the manufacture and optical characterization of an optical fiber nanotip coated with molecular beacons potentially capable to perform intracellular measurements. The optical characterization showed the good performance of the obtained nanotip in terms of selectivity and sensitivity, and a limit of detection of 0.57 nM was achieved. The treatment with HCl, besides allowing a regeneration of the nanotip, is able to provide a better performance of the molecular beacon with a further decrease of the limit of detection.


Education in diabetes mellitus is an ongoing process to promote/encourage the acquisition of knowledge on self-care skills and control of the disease and this process can be facilitated by social support from family. The literature shows the benefits of educational interventions for people with diabetes mellitus, however, there are few studies on its long-term effects. Objective: To assess the effectiveness of educational interventions focusing on social support after two yrs. of its completion.Quantitative/longitudinal study whose data before (T0) and after 12 months (T12) came from a clinical trial in which participants were given educational interventions through the “Conversation Maps in Diabetes”, tool and there was involvement of the family through an Intervention Group, done by phone calls. The third collection (T36) of data was from the population (N=164) that participated of clinical trial. The variables of interest was the knowledge of the disease/care, evaluated by “Diabetes Knowledge Scale” (DKN-A) validated in Brazil, and glycemic control assessed by glycated hemoglobin value obtained in electronic medical records. In the statistical analysis we used the nonparametric ANOVA for repeated measures.Of the 164 participants (82 of the Intervention and 82 of the Control Group), 95 (50 and 45 respectively) were interviewed. There were differences in the values of knowledge  and glycated hemoglobin  in the studied time (Figure The highest score of knowledge was at T12 in both groups, with a reduction in T36, reiterating the importance of education as an ongoing process. As for glycemic control, family social support seems to have influenced the maintenance of results.


Our histological results show complete incorporation of the quadriceps tendon into the host tissues, patellar survival and total integration of the allograft tibia, with relatively fewer osteocytes, into the host tibia. Vascularity and cellularity are reduced and delayed in the allograft but exhibit similar distributions to those in the autograft. The infrapatellar fat pad provides the main blood supply, and the lowest cellularity is observed in the patellar tendon close to the tibia in both the allograft and autograft. The biomechanical properties of the junction of quadriceps tendon and host tissues and those of the allograft patellar tendon are completely and considerably restored, respectively. Therefore, fresh-frozen CEM allograft reconstruction is viable, but the distal patellar tendon and the tibial block may be the weak links of the reconstruction. These findings provide new insight into the use of allograft in repairing disruption of the extensor mechanism.Different clinical results have been reported in the repair of extensor mechanism disruption using fresh-frozen complete extensor mechanism (CEM) allograft, creating a need for a better understanding of fresh-frozen CME allograft reconstruction. Here, we perform histological and biomechanical analyses of fresh-frozen CEM allograft or autograft reconstruction in an  Extensor mechanism disruption, either from severe fracture or tendon rupture, typically occurs after or during total knee arthroplasty (TKA), with an incidence of less than 2.5%1234578931112141516618192021224624252627282930Fresh-frozen CEM allograft reconstruction was first demonstrated in patellar tendon rupture after TKA4629271415161719in vivo rabbit model of fresh-frozen CEM allograft or autograft reconstruction and to investigate whether the allograft is a viable alternative for extensor mechanism reconstruction by conducting histological and biomechanical analyses during the first six months after surgery. A rabbit model of fresh-frozen CEM allograft or autograft reconstruction was first established. Quantitative analysis of cellularity and qualitative evaluation of vascularity were performed using Indian-ink perfusion and hematoxylin and eosin (H&E) staining. The strength of the junction of the quadriceps tendon and the soft tissues and the strength of the patellar tendon were measured using biomechanical test. We hypothesized that fresh-frozen CEM allograft reconstruction may be viable because the allograft would be incorporated into the host tissues even during a low remodeling process. Our study provides evidences for continuity between the surviving allograft and the surrounding tissues, despite a reduced number of osteocytes in the allograft tibia, the main blood supply from the infrapatellar fat pad, and significant restoration of the biomechanical properties of the allograft tendons, despite their low cellularity. These results indicate that fresh-frozen CEM allograft is a viable alternative for extensor mechanism reconstruction, but the distal patellar tendon and the tibial block may be the weak links of the reconstruction.This study was designed to establish an The 84 rabbits included in this study all survived the experiment. One specimen was excluded from the autograft group (n = 28) at 4 weeks postoperatively because of loose suture and infection. One specimen in the allograft group (n = 56) and one specimen in the autograft group were excluded at 12 weeks postoperatively due to improper performance of the biomechanical test.As shown in As shown in As shown in At 4 weeks, the central portion of the quadriceps tendon close to the patella  and the As shown in As shown in As shown in In the patellar tendon, almost all parameters were significantly lower at 4 weeks than those at 8, 12 and 24 weeks in both the allograft and autograft groups. Compared with those in the autograft group, almost all parameters in the allograft group were significantly lower at all time points. Compared with those in the control group, almost all parameters in both the allograft and autograft groups were significantly lower at all time points. In both the allograft and autograft groups, the cross-sectional area of the patellar tendon was significantly smaller at 24 weeks than at 4 weeks, but it was significantly larger than that in the control group at all time points. The maximum load to failure, stiffness at failure, maximum stress and elastic modulus in the allograft group at 24 weeks exhibited 73.6%, 69.5%, 65.8% and 63.9%, respectively, of those in the autograft group and 70.7%, 61.9%, 54% and 43% , respectively, of those in the control group. The maximum load to failure, stiffness at failure, maximum stress and elastic modulus in the autograft group at 24 weeks exhibited 89%, 82.9%, 77.1% and 62.1%, respectively, of those in the control group.Although different findings have been reported in the repair of extensor mechanism disruption using fresh-frozen CEM allograft, few, if any, studies have conducted histological and biomechanical analyses of the remodeling of the allograft. In this study, we chose rabbits to perform fresh-frozen CEM allograft or autograft reconstruction because remodeling of cortical bone in rabbits has been shown to be similar to that in humansin situ implantation of the autograft, resulting in the absence of a degeneration stage in the parts close to the infrapatellar fat pad.We show that the patterns of vascular and cellular distributions in the allograft patellar tendon in the sagittal and vertical axes are similar to those in the autograft patellar tendon, although they differ from those in the control. In the sagittal axis, the main blood supply for the patellar tendon in both the allograft and autograft groups is from the infrapatellar fat pad. This is in agreement with previous studies demonstrating that the vessels of the patellar tendon used for anterior cruciate ligament reconstruction were typically obtained from the infrapatellar fat pad surrounding the graft363736373839404142i.e., thin layer of chondrocytes and large spaces between the trabeculae) in the allograft patella, revascularization and the increased number of chondrocytes indicate that the patella is survived. The complete integration of the allograft tibia into the host tibia is similar to the findings of some previous studies which found the healing of the allograft bone block to the host tissue at all tibial junctions1415161719Although the appearance of abrasion and osteoporosis  were used in this experiment. Each rabbit was checked for general health, housed in commercial animal cages (49 cm × 35 cm × 32 cm) and had free access to food and water. The experimental protocol was approved by the Animal Care and Use Committee of the Fourth Military Medical University. The methods were carried out in accordance with the approved guidelines. Every effort was made to improve the comfort of the rabbits.The rabbits were divided into the allograft group (n = 56) and the autograft group (n = 28). In the allograft group, 28 rabbits were used to harvest right CEMs. The 28 right CEMs were soaked in gentamicin solution  and immediately stored in sterile bags at −80 °C  for one week. Each of the remaining 28 rabbits received one right fresh-frozen CEM allograft. In the autograft group, 28 rabbits underwent right CEM autograft reconstruction. At 4, 8, 12 and 24 weeks postoperatively, 2 rabbits were used from each group for Indian-ink perfusion and histological analysis, and 5 rabbits were used for biomechanical test. From each rabbit, both CEMs were harvested, and the left CEM served as the control group. The left CEMs frozen at −80 °C for 7 days served as the −80 °C group. Postoperatively, the rabbits were returned to their own cages, softly touched every day, and given an intramuscular injection of cefazolin sodium (0.25 g) for 7 consecutive days. No analgesics were administered. The cast was removed after 3 weeks of immobilization.2S under anesthesia . A median knee incision was performed and the quadriceps tendon was first severed 1.5 cm above the patella. The soft tissues alongside the patella and the patellar tendon were then separated. Finally, the CEM was harvested following the creation of a tibial bone block that was 1.2 cm long, 0.5 cm wide and 0.4 cm thick. Before implantation, the fresh-frozen CEM allograft stored in a sterile bag was bathed in 37 °C water for 5 minutes, and 2 number-5 nonabsorbable sutures [Johnson & Johnson Medical (China) Ltd.] were placed along the lateral and medial aspects of quadriceps tendon in a modified Krackow fashionin situ  before depilation of the abdomen and the legs. Subsequently, a median abdominal incision was created, followed by the exposure of the abdominal aorta and the inferior vena cava and the ligation of their branches. Before a syringe needle was inserted into the distal of the abdominal aorta, the proximal of the abdominal aorta and the inferior vena cava were tied off. The syringe needle was secured in the vessel, and the distal of the inferior vena cava was transected. Approximately 500 mL of heparinized saline (25000 IU/L) was needed to perfuse the abdominal aorta at a rate of 4 ml/min until the blood outflow from the distal of the inferior vena cava stopped and the fluid became clear. Next, a 10% formalin solution and a 10% Indian-ink solution  filtered through quantitative filter paper  were serially perfused for 30 minutes each at the same rate as the saline perfusion. At the end of the India-ink perfusion, the distal of the inferior vena cava was tied off. An angiograph of the left CEM is shown in The Indian-ink perfused CEM, including the proximal and distal of the adjacent tissues, was first fixed in Bouin’s fluid for 24 hours. Subsequently, it was divided into 5 parts after being transected along the midline of the patella and tibia block: 1) the junction of the quadriceps tendon and the soft tissues, the quadriceps tendon and half of the patella; 2) the upper part of the patellar tendon and half of the patella; 3) the middle part of the patellar tendon; 4) the lower part of the patellar tendon and half of the tibial block; 5) the tibial block insertion site. The specimens, excluding the middle part of the patellar tendon, were decalcified in 10% ethylenediaminetetraacetic acid for 4 weeks. Next, they were dehydrated in a graded ethanol solutions and embedded in paraffin. Five sections with a thickness of 5 μm, spaced at an interval of 50 μm, were obtained from each specimen , stained with H&E and evaluated by 2 of the authors (G.C. and H.Z.) in a blinded manner.2) in each section was identified and the number of cells was counted. At the distal of the upper part  of the patellar tendon, one region of interest (0.5651 mm2) from the dorsal, central and ventral portions of each section was analyzed, and the cellularity of the dorsal, central and ventral patellar tendons was calculated. In addition, the cellularity of the central portion of the quadriceps tendon close to the patella was calculated. Histological remodeling processes of the junction of the quadriceps tendon and the soft tissues, the patella and the tibial block insertion site were described.All of the sections were observed at different magnifications for the quantitative and qualitative analyses of cellularity and vascularity using a high-resolution microscope . The cellularity of the quadriceps tendon and the patellar tendon were evaluated in longitudinal and median section. The vascularity of the middle part of the patellar tendon was assessed in transverse section. Each section was divided into a ventral portion, a central portion and a dorsal portion. In the central portions of the upper part of the patellar tendon close to the patella and the lower part of the patellar tendon close to the tibia, one region of interest .


Carnegiea gigantea) in the Cactus Forest of the Rincon Mountain District of Saguaro National Park near Tucson, AZ. On 6 four-hectare plots, each saguaro was censused and a methodical search for new saguaros was conducted annually each year from 1942 through 2016, with the exception of 1955. Regeneration has been episodic with 828 plants established from 1959 through 1993 compared with 34 plants established between 1942 and 1958 and only three plants established after 1993. The years preceding 1959 and following 1993, include some of the driest decades in centuries in southern Arizona. While woodcutting and cattle grazing are believed to be among the causes of decades of failed regeneration prior to 1958, neither of these factors contributed to the failed regeneration following 1993. The height structure of the population from 1942 to 2016 shifted dramatically from a population dominated by large saguaros  in the first three decades of the study to a population dominated by small saguaros  in the most recent two decades. Mortality is shown to be strongly age dependent. In the year following the 2011 catastrophic freeze, 21 of 59 plants older than 80 years died compared with zero deaths in 270 plants between the ages of 29 and 80 years. Saguaros under 40 years old, growing under small shrubs or in the open, have a lower probability of survival than better protected saguaros. Long-term population monitoring is essential to understanding the complex impacts of human and environmental factors on the population dynamics of long-lived species.Annual census data spanning seventy-five years document mortality and regeneration in a population of saguaro cactus ( Carnegiea gigantea [Engelm.] Britton & Rose) at the Carnegie Institution Desert Laboratory on Tumamoc Hill in Tucson, Arizona [Beginning with Forrest Shreve’s classic studies of the giant saguaro cactus  section of the Cactus Forest devoted to researching saguaro mortality . In addiSaguaros begin to produce fruit at 30 to 45 years of age and have a lifespan estimated to be between 125 to 175 years . The amoBecause the saguaro is an icon of both the Sonoran Desert and the state of Arizona, there is considerable popular interest in the health of the saguaro population in the face of changing climatic conditions. The current report represents a detailed study of saguaro mortality and regeneration for a period of seventy-five years that includes one episode of regeneration spanning approximately three decades in one part of the Cactus Forest of the Rincon Mountain District of Saguaro National Park.This study was carried out in the Rincon Mountain District of Saguaro National Park in Tucson, Arizona with the permission of the National Park Service. No samples were collected. The study was done in accordance with the conditions set forth in our research permit from the National Park Service. The current permit number is SAGU-2014-SCI-0001.Between October 1940 and May 1941, University of Arizona engineering students surveyed the corners of sixty-four 4-hectare plots forming a grid in Section 17, Township 14 South, Range 16 East, at Saguaro National Monument  east of Tucson, Arizona . A numbeIn any study spanning over seven decades, events will occur adding to the individuality of plots and this study is no exception. As noted above, three plots were “treated” in 1942, whereas three plots were not. The north half of section 17 (untreated) was privately owned until the 1970s, when it was purchased by the Monument. Cattle grazing occurred on all plots until the mid-1960s when a boundary fence excluded grazing from the southern plots. Grazing was excluded from the northern plots later. Saguaros were transplanted into one plot in 1958 [The saguaro population discussed here is located in the flats of the Rincon Mountain District of the Saguaro National Park. Steenbergh and Lowe ,18 emphaParkinsonia microphylla Torr.) and some mesquite (Prosopis velutina Woot.) whereas wash bottoms are dominated by mesquite, creosote bush (Larrea tridentata (DC) Coville), and ribbons of catclaw acacia  greggii A. Gray) along watercourses, but contain few saguaro and paloverde.The flats where the plots are located consist of quaternary alluvial soil of the lower bajada of the Rincon Mountains. The landscape slopes downward gradually from south to north and is interspersed with a network of northwardly-flowing washes that are usually 2–4 m below grade and of varying widths . The resThe annual saguaro census consists of visiting each saguaro that was living in the previous year, assessing its condition, and noting mortality. In 1942, all plants were classified into five height classes: class I (0.0–1.8 m); class II (1.8–3.7 m); class III (3.7–5.5 m); class IV (5.5–7.3 m) and class V (>7.3 m). Plants were classified by height class again in 1960, and in many subsequent years. The height classes of plants were evaluated in several ways over the course of the study. In most years, when a plant appeared to be near a change in height class, we evaluated the height class by sighting from a distance using a 1.8 m reference standard placed next to the saguaro. A clinometer was used in 1983 and 1987 to measure all plants and check height classes. All plants were measured in 2011 with a fiberglass extension pole. Since 1968, we have counted the number of branches greater than approximately 10 cm diameter on each plant. Individual plants are identified based on their map location, height or height class, and individual marks .Each spring, typically in March through 1999 and in January through March thereafter, we methodically search for new plants. When found, we map them, measure them, describe any marks or scars, and note the nurse plant and the direction of the saguaro from the base of the nurse plant. Care is taken with descriptions and measurements of individuals to insure that newly discovered plants are found in subsequent years. In the 1990s the original maps were digitized and entered into an ArcInfo geographic information system (GIS), which has been maintained using Arc View 3.1 . After 1979, except for 1982, the height of every plant less than 1.8 m tall was measured to the nearest 0.5 cm every year.We estimated the year of germination for new plants found in our plots based on an estimate of the age of the plant in the year it was found. The standard way to estimate age of saguaros is to use a table relating height to age developed from growth measurements taken of plants of various heights ,7,8,19. To estimate age from height requires a three-step process. First, develop a model of growth from height (we use log(growth) from log(height)) . Then, uBecause of this variability in growth, the sooner a saguaro is discovered, the better the age estimate. In a natural setting, saguaros can take several years to reach the height of 2 cm (Table 2–2 in ), so thePierson, Turner, and Betancourt  describeMortality was analyzed in two ways: For the saguaros counted in the study plots in 1942, survival curves were calculated for cohorts defined by their height class in 1942. Survival curves for these plants show the percent of each 1942 cohort that survived each year from 1942 to the present. For saguaros found after 1942, we have a better estimate of age based on the height of the saguaro in the year it was found. So, we have good age estimates for each of these saguaros from the time of discovery until the time of death. Annual survival/mortality observations of 867 saguaros found in the plots after 1960 were used to construct survival curves for saguaros between the ages of 10 years and 40 years for each of three cover classes . The datThe structure of the saguaro population in this study has been shaped by alternating periods of regeneration success or failure combined with ongoing mortality of older plants. The impact of this dynamic on the height/age structure of the population over the decades has been dramatic . The popThe number of plants established in each year from 1937 to 2002 was examined using all plants found in the study area after 1942 . Few of There was a significant reduction in the number of plants establishing after 1984  droppingReproductive potential, measured as the number of stem tips/hectare, depends on the height/age structure of the population, which has changed dramatically over the 75 years . The decThe mortality of the 1942 population was strongly dependent on the height (age) of the plants in 1942 . For exaWe do not have good age estimates for the height classes in 1942. For an approximation, we can use the median ages of plants in those height classes in 2015. We have fairly precise age estimates for plants discovered after 1958 based on their height at the time of discovery. The median age of plants in 2015 in height classes V, IV, III, II, and I is >85, 73, 51, 41, and 34 years respectively. Note that faster growing, and thus, younger plants will enter the taller height classes first and be followed by the older plants. Therefore, height class III, in particular, could contain a higher proportion of younger faster growing plants from the episode of regeneration (1959 to 1993) than height class III plants in, for example, a decade from now. So, the median age estimate for the height class III in 1942, based on the current height class III ages may somewhat low.Adding the median survival time  to an esnd and 3rd, 2011 there was a freeze in the Tucson area that met the Bower’s criteria. In the 2012 census, following the 2011 freeze, there was a sharp drop in the percent survival of the 1942 height class I plants   large trees , C2) small trees and large shrubs, and C3) small shrubs, grasses, and plants that provide minimal cover, or no cover plant at all . Five of< 0.001) .2 = 155, df = 2; p < 0.001). Overall, mortality of small saguaros in cover class C3 (62%) is significantly higher than mortality in cover class C1 (31%) and cover class C2 (21%)  for saguaros that germinated during the episode of population regeneration  ,7, the PBetancourt et al.  state thth century, whereas in our plots, the large number of saguaros in height class III in 1942 suggests the late 19th century was a particularly good period of regeneration based on the following method. Our estimate of the age range for height class III plants is younger and longer than an estimate based on deriving the age from the limits of the height class (3.6 to 5.4 m) alone. We base our age estimates on the height when the plant is found and follow its growth until it reaches that height class. Rapidly growing individuals reach height class III by age 38 years and the oldest member of 2015 height class III is 69 years old. Therefore, our estimate of the age of plants in height class III in 2015 range between 38 and 69 years old or roughly 40 to 70 years old. Using the Steenbergh and Lowe estimates for saguaros 3.6 m and 5.4 m tall (the height class limits) gives a narrower age range of 53 to 68 years [The contrast between our population and the population at Tumamoc Hill further illustrates the point that multiple local factors beyond broad regional patterns influence saguaro population regeneration. Pierson and Turner  reported class 3. to 5.4 mOne of the ten plots studied by Pierson, Turner, and Betancourt  is in thPierson, Turner, and Betancourt  suggest We have described in the methods section the history of the plots. There are not sufficient controls or replications to assess the impact of the differences in plot history on regeneration or mortality, so we have not analyzed the plots separately. No obvious effects stand out. Subjectively, the impact of microhabitat factors such as soil type, topography , and cover plant composition  are the primary influences on the patterns of regeneration and survival that we have observed, both within and between plots. The expansion of the road and picnic area into relatively good quality microhabitat could have influenced regeneration in one plot marginally.Weiss, Castro, and Overpeck  contrastIt is widely accepted that a major cause of saguaro mortality is catastrophic freeze  and thatErwinia cacticida [There were no catastrophic freezes from 1979 through 2011, over 30 years, and yet age-related mortality continued apace . One wayacticida , a bacteJohnson and Coppola  report tnd and 3rd with a high temperature of only 3.3 C on February 3rd), which is typical of a catastrophic freeze event [The 2011 freeze event was characterized by an extended period of sub-freezing temperatures Click here for additional data file.S1 TableThe coefficients were obtained by stepwise linear regression to model log growth (y) from log height (x) for saguaros between the ages of 8 years and 30 years.(DOCX)Click here for additional data file.S2 TableThese models are used to determine the age of saguaros based on their height at the time of discovery.(DOCX)Click here for additional data file.