Background General dental practitioners provide the majority of endodontic treatment in Jordan. The aim of this study was to gather information on the methods, materials and attitudes employed in root canal treatment by dentists in North Jordan, in order to evaluate and improve the quality of current practice.
Background
General dental practitioners provide the majority of endodontic treatment in Jordan. The aim of this study was to gather information on the methods, materials and attitudes employed in root canal treatment by dentists in North Jordan, in order to evaluate and improve the quality of current practice.Methods A questionnaire was posted to all registered general dental practitioners working in private practice in Irbid Governate in North Jordan (n = 181). The questionnaire included information on methods, materials and techniques used in endodontic treatment.
Methods
A questionnaire was posted to all registered general dental practitioners working in private practice in Irbid Governate in North Jordan (n = 181). The questionnaire included information on methods, materials and techniques used in endodontic treatment.Results Reply rate was 72% (n = 131). The results demonstrated that only five dentists used rubber dam occasionally and not routinely. The majority used cotton rolls for isolation solely or in combination with a high volume saliva ejector (n = 116). The most widely used irrigants were sodium hypochlorite and hydrogen peroxide, which were used by 32.9% (n = 43) and 33.6% (n = 44) of the respondents, respectively. Forty eight percent of the respondents (n = 61) used the cold lateral condensation technique for canal obturation, 31.3% (n = 41) used single cone, 9.9% (n = 13) used vertical condensation and 12.2% (n = 16) used paste or cement only for the obturation. The majority used zinc oxide eugenol as a sealer (72.5%). All, but one, respondents used hand instruments for canal preparation and the technique of choice was step back (52.7%). More than 50% (n = 70) of the dentists took one radiograph for determining the working length, whilst 22.9% (n = 30) did not take any radiograph at all. Most practitioners performed treatment in three visits for teeth with two or more root canals, and in two visits for teeth with a single root canal.
Results
Reply rate was 72% (n = 131). The results demonstrated that only five dentists used rubber dam occasionally and not routinely. The majority used cotton rolls for isolation solely or in combination with a high volume saliva ejector (n = 116). The most widely used irrigants were sodium hypochlorite and hydrogen peroxide, which were used by 32.9% (n = 43) and 33.6% (n = 44) of the respondents, respectively. Forty eight percent of the respondents (n = 61) used the cold lateral condensation technique for canal obturation, 31.3% (n = 41) used single cone, 9.9% (n = 13) used vertical condensation and 12.2% (n = 16) used paste or cement only for the obturation. The majority used zinc oxide eugenol as a sealer (72.5%). All, but one, respondents used hand instruments for canal preparation and the technique of choice was step back (52.7%). More than 50% (n = 70) of the dentists took one radiograph for determining the working length, whilst 22.9% (n = 30) did not take any radiograph at all. Most practitioners performed treatment in three visits for teeth with two or more root canals, and in two visits for teeth with a single root canal.Conclusions This study indicates that dentists practicing in North Jordan do not comply with international quality standards and do not use recently introduced techniques. Many clinicians never take a radiograph for determining the working length and never used rubber dam or intra-canal medicaments.
Conclusions
This study indicates that dentists practicing in North Jordan do not comply with international quality standards and do not use recently introduced techniques. Many clinicians never take a radiograph for determining the working length and never used rubber dam or intra-canal medicaments.

Background Root canal treatment is considered an essential element in the dental services provided to the population in developed countries. Various investigations were, therefore, carried out to explore the standard of root canal treatment carried out by general dental practitioners in Europe [ 1 - 3 ]. It is the responsibility of the academics and dental schools to prepare their students to adopt the guidelines and recommended standards in root canal debridement, shaping and obturation [ 4 , 5 ]. Several studies have revealed that the majority of dentists do not comply with the formulated guidelines on the quality of root canal treatment [ 1 - 3 , 6 ]. These studies investigated the attitude of dentists in Western countries such as Germany [ 1 ], UK [ 2 ], Belgium [ 3 ] and the USA [ 6 ]. On the other hand, few studies have investigated the attitude of general dental practitioners toward various aspects of endodontic treatment in developing countries [ 7 - 9 ]. The majority of endodontic treatment in Jordan is provided by the general dental practitioners due to absence of specialists in endodontics and to the lack of postgraduate programs in Jordan. The purpose of the current study was, therefore, to investigate the attitude of dentists toward endodontic treatment and to explore the materials and methods employed by general dental practitioners in North Jordan and to compare these findings with well-acknowledged international academic standards.
Background
Root canal treatment is considered an essential element in the dental services provided to the population in developed countries. Various investigations were, therefore, carried out to explore the standard of root canal treatment carried out by general dental practitioners in Europe [ 1 - 3 ].It is the responsibility of the academics and dental schools to prepare their students to adopt the guidelines and recommended standards in root canal debridement, shaping and obturation [ 4 , 5 ]. Several studies have revealed that the majority of dentists do not comply with the formulated guidelines on the quality of root canal treatment [ 1 - 3 , 6 ]. These studies investigated the attitude of dentists in Western countries such as Germany [ 1 ], UK [ 2 ], Belgium [ 3 ] and the USA [ 6 ]. On the other hand, few studies have investigated the attitude of general dental practitioners toward various aspects of endodontic treatment in developing countries [ 7 - 9 ].The majority of endodontic treatment in Jordan is provided by the general dental practitioners due to absence of specialists in endodontics and to the lack of postgraduate programs in Jordan. The purpose of the current study was, therefore, to investigate the attitude of dentists toward endodontic treatment and to explore the materials and methods employed by general dental practitioners in North Jordan and to compare these findings with well-acknowledged international academic standards.Methods A postal survey of general dental practitioners in Jordan was carried out to investigate common materials and methods employed in root canal treatment. A questionnaire was developed and piloted by sending it to 20 general dental practitioners. According to the replies, the questionnaire was modified. Few questions were added and others were reworded. Additionally, the questionnaire was provided in Arabic and English language. The finally modified questionnaire was posted to all registered general dental practitioners (n = 181) listed in the records of the Jordanian Dental Association and working in private practices in Irbid Governate in North Jordan, 'Questionnaire [see  Additional file 1 ]'. The questionnaire consisted of 28 questions concerning different aspects of endodontic treatment including the provision of molar endodontics, root canal therapy stages, materials, the choice of instruments, the use of rubber dam and isolation methods, number of appointments, number of radiographs taken throughout the treatment, the use of canal irrigants, the use of intracanal medicaments, the choice of obturation technique, temporary and permanent coronal restoration, and case monitoring and follow-up. There was a space made available in the questionnaire for free comments of respondents. The questionnaire was accompanied by an explanatory covering letter. To investigate the influence of the years of practical experience on the materials and techniques employed, the sample was divided into groups based on the years of professional experience: group 1, up to 5 years; group 2, 6–10 years; group 3, 11–15 years; group 4, 16–20 years, and group 5, more than 20 years. The collected data were entered into a personal computer and analyzed using the statistical package SPSS. Simple descriptive statistics were used together with Chi-square (χ 2 ) test. The chosen level of significance was set at  P  < 0.05. Unanswered questions were treated as missing values.
Methods
A postal survey of general dental practitioners in Jordan was carried out to investigate common materials and methods employed in root canal treatment. A questionnaire was developed and piloted by sending it to 20 general dental practitioners. According to the replies, the questionnaire was modified. Few questions were added and others were reworded. Additionally, the questionnaire was provided in Arabic and English language. The finally modified questionnaire was posted to all registered general dental practitioners (n = 181) listed in the records of the Jordanian Dental Association and working in private practices in Irbid Governate in North Jordan, 'Questionnaire [see  Additional file 1 ]'. The questionnaire consisted of 28 questions concerning different aspects of endodontic treatment including the provision of molar endodontics, root canal therapy stages, materials, the choice of instruments, the use of rubber dam and isolation methods, number of appointments, number of radiographs taken throughout the treatment, the use of canal irrigants, the use of intracanal medicaments, the choice of obturation technique, temporary and permanent coronal restoration, and case monitoring and follow-up. There was a space made available in the questionnaire for free comments of respondents. The questionnaire was accompanied by an explanatory covering letter.To investigate the influence of the years of practical experience on the materials and techniques employed, the sample was divided into groups based on the years of professional experience: group 1, up to 5 years; group 2, 6–10 years; group 3, 11–15 years; group 4, 16–20 years, and group 5, more than 20 years. The collected data were entered into a personal computer and analyzed using the statistical package SPSS. Simple descriptive statistics were used together with Chi-square (χ 2 ) test. The chosen level of significance was set at  P  < 0.05. Unanswered questions were treated as missing values.PResults Of the 181 questionnaires distributed, 131 completed replies were received, which is a 72% response rate. The high response rate ensured that this study was representative for the general dental practitioners in North Jordan. All the respondents performed endodontic treatment including molar teeth. However, none of the dentists reported that they would refer patients for a specialised endodontist opinion except cases, which were difficult or did not respond to initial treatment provided. The distribution of the repondents according to the years of professional experience is shown in Table  1 . Years in practice were not evenly distributed amongst the total respondents. The number of the first two groups (0–5 and 6–10) consisted of more than half the total respondents due to the significant increase in the number of graduates in the last 10 years. Seventy four percent of the respondents were males, 26% were females. These findings are consistent with the statistics obtained from the Jordanian Dental Association. In the current study, no statistically significant differences were found between the different periods of professional experience and any of the materials, instruments or techniques employed ( P  > 0.05). Table 1 Data related to professional experience of the respondents. Years of Professional experience Frequency Percentage % 0–5 43 32.8 6–10 28 21.4 11–15 26 19.8 16–20 20 15.3 >20 14 10.7 Table  2  shows the hand instruments used for preparation of the root canal. K-files were the most popular instruments. Root canal preparation was performed using K-files solely (30.5%) or in combination with other instruments (93.1%). Only one practitioner reported using engine-driven instruments (Profile, Dentsply Maillefer, ballaigues, Switzerland). Table 2 The choice of root-canal preparation techniques and instruments Root canal preparation techniques Root canal instrument Technique Frequency % Instrument Frequency % Filing (push-pull) 36 27.5 File 40 30.5 Step back 69 52.7 Reamer 3 0.2 Step down 26 19.8 Hedström file 6 4.6 File + hedström file 30 22.9 File + reamer 20 15.3 File, reamer + hedström file 32 24.4 The majority of dentists instrumented the canal using the step back technique. The next most popular preparation technique was the filing (push-pull) technique followed by the step down technique (Table  2 ). The vast majority used gutta-percha points as their priniciple root filling material (87.8%), whilst 12.2% reported using only paste or cement to obturate the canal. Cold lateral compaction was the most common obturation technique (Table  3 ). The majority of dentists reported the use of a zinc oxide based sealer with the gutta-percha points (72.5%) followed by a calcium hydroxide based sealer, Sealapex (13.7%) (Table  3 ). Few dentists (n = 8) used the sealer Endomethasone as a paste root canal filling. Table 3 The choice of obturation technique and type of sealer. Root canal obturation techniques Type of sealer Technique Frequency % Type of material Frequency % Single cone 41 31.3 Zinc oxide-eugenol 95 72.5 Lateral condensation 61 46.6 Sealapex 18 13.7 Vertical condensation 13 9.9 Endomethason 10 7.6 Cement only 16 12.2 Other 8 6.2 Intracanal medication was used by 63% of the respondents. The most common material used was tricresol formalin followed by calcium hydroxide. Other formulations were also used (Table  4 ). Table 4 The frequency and percentages of intracanal medications used. Type of product Frequency Percentage % Calcium hydroxide 15 11.5 Formaldehyde 6 4.6 Tricresol formaline 45 34.4 Dexamethasone 1 0.8 Iodophorm 5 3.8 CMCP * 2 1.5 Other 8 6.1 None 49 37 * Camphorated monochlorophenol Sodium hypochlorite and hydrogen peroxide solutions were used equally as an irrigating solutions. The most popular concentration of sodium hypochlorite was 3% which was used by 14.5% (n = 19) of the repondents, with only 2.3% (n = 3) using a 0.5% concentration. The most commonly used concentration of hydrogen peroxide was 3%, which was used by 21.4% (n = 28) of the respondents. The remainder used either normal saline or local anesthetic solutions (Table  5 ). Table 5 Data related to the choice of root-canal irrigants. Root-canal irrigants used Concentration of NaOCl used Concentration of H 2 O 2  used Type Frequency % Concentration (%) Frequency % Concentration (%) Frequency % Sodium hypochlorite 43 32.8 0.5 3 2.3 1 2 1.5 Normal saline 32 24.4 1 3 2.3 2 2 1.5 Hydrogen peroxide 44 33.6 2 8 6.1 3 28 21.4 Local anesthetic solution 2 1.5 3 19 14.5 4 2 1.5 None 10 7.6 5 3 2.3 6 10 7.6 6 5 3.8 Do not use H 2 O 2 87 66.4 Do not use NaOCl 88 67.2 None of the dentists reported using rubber dam routinely to isolate the field of operation during root canal therapy. However, only five dentists reported using rubber dam occasionally but not as a routine practice. The majority of the general dental practitioners used cotton rolls solely (n = 68) or cotton rolls in combination with a high volume saliva ejector (n = 116) to reduce contamination with saliva (Figure  1 ). Figure 1 The number of dentists using different isolation methods. The number of visits required to complete root canal treatment related to the number of root canals in a tooth is shown in Figure  2 . It demonstrates that general dental practitioners complete root canal treatment in more than two visits for teeth with two or more root canals. However, half the respondents (49.7%) reported completing root canal treatment for teeth with single root canal in two visits. Figure 2 The number of visits according to the number of root canals per tooth. Twenty seven percent of the practitioners took 3 radiographs for routine root canal treatment. 22.9% took only 2 radiographs. However, 23% reported taking only one preoperative radiograph with 4% taking only one radiograph for determining the working length. The remaining 22.9% of respondents undertook root canal treatment without taking any radiograph. Only 14.5% of the respondents reported monitoring the root treated tooth radiographically after a period of 6 months. However, many of them mentioned that they would take a follow-up radiograph only if patients could afford to pay for it. The remainder indicated that they do not monitor their patients mostly for financial reasons and that patients would not return for follow-up appointment unless they have postoperative symptoms. Zinc oxide eugenol cement was the most commonly placed temporary filling (92%). All dentists reported using amalgam for posterior teeth and composite for anterior teeth as a permanent coronal restorative material. All practitioners completed the restorations themselves. Sixty four percent of the respondents preferred to wait from 1 to 2 weeks after obturation before placing the permanent coronal filling, whilst the remainder placed the restoration immediately after completion of the treatment.
Results
Of the 181 questionnaires distributed, 131 completed replies were received, which is a 72% response rate. The high response rate ensured that this study was representative for the general dental practitioners in North Jordan.All the respondents performed endodontic treatment including molar teeth. However, none of the dentists reported that they would refer patients for a specialised endodontist opinion except cases, which were difficult or did not respond to initial treatment provided.The distribution of the repondents according to the years of professional experience is shown in Table  1 . Years in practice were not evenly distributed amongst the total respondents. The number of the first two groups (0–5 and 6–10) consisted of more than half the total respondents due to the significant increase in the number of graduates in the last 10 years. Seventy four percent of the respondents were males, 26% were females. These findings are consistent with the statistics obtained from the Jordanian Dental Association. In the current study, no statistically significant differences were found between the different periods of professional experience and any of the materials, instruments or techniques employed ( P  > 0.05).PData related to professional experience of the respondents.Years of Professional experienceYears of Professional experienceTable  2  shows the hand instruments used for preparation of the root canal. K-files were the most popular instruments. Root canal preparation was performed using K-files solely (30.5%) or in combination with other instruments (93.1%). Only one practitioner reported using engine-driven instruments (Profile, Dentsply Maillefer, ballaigues, Switzerland).The choice of root-canal preparation techniques and instrumentsRoot canal preparation techniquesRoot canal preparation techniquesRoot canal instrumentRoot canal instrumentThe majority of dentists instrumented the canal using the step back technique. The next most popular preparation technique was the filing (push-pull) technique followed by the step down technique (Table  2 ).The vast majority used gutta-percha points as their priniciple root filling material (87.8%), whilst 12.2% reported using only paste or cement to obturate the canal. Cold lateral compaction was the most common obturation technique (Table  3 ). The majority of dentists reported the use of a zinc oxide based sealer with the gutta-percha points (72.5%) followed by a calcium hydroxide based sealer, Sealapex (13.7%) (Table  3 ). Few dentists (n = 8) used the sealer Endomethasone as a paste root canal filling.The choice of obturation technique and type of sealer.Root canal obturation techniquesRoot canal obturation techniquesType of sealerType of sealerIntracanal medication was used by 63% of the respondents. The most common material used was tricresol formalin followed by calcium hydroxide. Other formulations were also used (Table  4 ).The frequency and percentages of intracanal medications used.Type of productType of product* Camphorated monochlorophenolSodium hypochlorite and hydrogen peroxide solutions were used equally as an irrigating solutions. The most popular concentration of sodium hypochlorite was 3% which was used by 14.5% (n = 19) of the repondents, with only 2.3% (n = 3) using a 0.5% concentration. The most commonly used concentration of hydrogen peroxide was 3%, which was used by 21.4% (n = 28) of the respondents. The remainder used either normal saline or local anesthetic solutions (Table  5 ).Data related to the choice of root-canal irrigants.None of the dentists reported using rubber dam routinely to isolate the field of operation during root canal therapy. However, only five dentists reported using rubber dam occasionally but not as a routine practice. The majority of the general dental practitioners used cotton rolls solely (n = 68) or cotton rolls in combination with a high volume saliva ejector (n = 116) to reduce contamination with saliva (Figure  1 ).The number of visits required to complete root canal treatment related to the number of root canals in a tooth is shown in Figure  2 . It demonstrates that general dental practitioners complete root canal treatment in more than two visits for teeth with two or more root canals. However, half the respondents (49.7%) reported completing root canal treatment for teeth with single root canal in two visits.Twenty seven percent of the practitioners took 3 radiographs for routine root canal treatment. 22.9% took only 2 radiographs. However, 23% reported taking only one preoperative radiograph with 4% taking only one radiograph for determining the working length. The remaining 22.9% of respondents undertook root canal treatment without taking any radiograph.Only 14.5% of the respondents reported monitoring the root treated tooth radiographically after a period of 6 months. However, many of them mentioned that they would take a follow-up radiograph only if patients could afford to pay for it. The remainder indicated that they do not monitor their patients mostly for financial reasons and that patients would not return for follow-up appointment unless they have postoperative symptoms.Zinc oxide eugenol cement was the most commonly placed temporary filling (92%). All dentists reported using amalgam for posterior teeth and composite for anterior teeth as a permanent coronal restorative material. All practitioners completed the restorations themselves. Sixty four percent of the respondents preferred to wait from 1 to 2 weeks after obturation before placing the permanent coronal filling, whilst the remainder placed the restoration immediately after completion of the treatment.Discussion The response rate was 72%. It was higher than in many previous surveys conducted in Western countries with better communication infrastructure [ 2 , 7 , 10 , 11 ]. The vast majority of the respondents did not practice single visit root canal treatment. This finding was in agreement with the results of a previous study undertaken in another developing country, Sudan [ 9 ]. However, a study from the US [ 12 ] demonstrated a clear inclination to single visit endodontics, especially in cases without apical periodontitis. Single visit treatment appears to have gained more popularity and an increased credibility in the pre-clinical endodontic teaching in America and Europe [ 4 ]. Another survey [ 3 ], showed that a high percentage of Flemish dentists performed single visit root canal treatment. Multiple visit endodontic treatment could be a direct result of lacking adequate clinical time to complete the treatment in a single visit. The dentists may prefer to wait till the complete subsidence of pain and other symptoms before obturating the canal system. Another possible explanation could be that the initial visit was spent for treating the pain and acute symptoms [ 3 ]. Although the application of rubber dam is always recommended as a standard during root canal treatment procedure to provide isolation, protection and improve visual access, only five dentists reported using rubber dam very occasionally and not as a routine practice. Similar findings were found in Sudan (2%) and among Flemish dentists (3.4%) [ 6 , 8 ]. However, 59% of American dentists [ 6 ], 60% of dentists in UK [ 13 ] and 57% of general dental practitioners in New Zealand [ 14 ] reported using rubber dam routinely in endodontic treatment. The reasons for not using rubber dam could be the extra cost, additional time, lack of adequate skills or training, absence of patient's acceptability or inadequate education in the undergraduate teaching curriculum. It was found that continuing education course attendees seem to be encouraged to use rubber dam [ 14 ]. In the current survey, most general dental practitioners used hydrogen peroxide and sodium hypochlorite solutions as canal irrigants. The same result was demonstrated amongst dentists in Switzerland [ 11 ]. Sodium hypochlorite is recommended as the material of choice for irrigating the root canal system because of its effective antimicrobial and tissue solving action [ 15 ]. The selection of irrigant could be associated with the use of rubber dam, as it was found that 70% of rubber dam users among British dentists irrigated with sodium hypochlorite, whilst non-users tended to use local anesthetic solution [ 13 ]. The current findings do not mirror these findings. The vast majority of our respondents were non-users of rubber dam and one third of them use sodium hypochlorite routinely. A similar trend toward using sodium hypochlorite as an irrigant despite not using rubber dam for isolation, was noticed amongst Flemish dentists [ 16 ]. In the UK, the majority of dentists used local anesthetic solution to irrigate the canal space [ 2 ]. The use of either sodium hypochlorite or hydrogen peroxide without isolating the field of operation tightly with a rubber dam presents an obviously hazardous practice in the use of potentially irritant irrigation solutions. Despite the fact that calcium hydroxide is recognized as the standard intracanal medicament for inter-appointment dressing [ 17 ], it was used by only 11.5% of the respondents. More than one third of the general practitioners reported using formaldehyde-containing materials. This finding is consistent with previous findings recorded for Sudanese dentists [ 9 ]. Although formaldehyde-containing products have been used for their antimicrobial and fixative properties, they are toxic to periradicular tissues [ 18 ] and may have mutagenic and carcinogenic potential [ 19 ]. The use of calcium hydroxide, as intracanal medication, should be encouraged among dentists in developing countries such as Jordan, as it is effective against most root canal pathogens and able to denature bacterial endotoxins [ 20 , 21 ]. It has, also, been reported to be the material of choice by dentists in the Western world [ 11 , 22 ]. The step back technique was the most popular canal preparation technique among North Jordanian general dental practitioners. The filing (push-pull) technique, on the other hand, was used by 27.5% of the respondents. In another study, 60.4% of Flemish dentists used the standard filing technique [ 16 ]. Generally, dentists in Jordan tended to use hand instruments and were not inclined to use more advanced engine driven techniques for shaping the root canal system. Almost half of the general dental practioners in North Jordan used cold lateral compaction of gutta-percha to obturate the root canal space. This technique is acknowledged universally and is the most common obturation technique [ 4 ]. However, 31.3% of the dentists in the current survey used a single cone technique, in common with 68% of Swiss dentists [ 11 ]. Additionally, 12.2% of respondents used only paste to obturate the root canal system. Seemingly, dentists in North Jordan are not strong advocates of the more recently introduced advanced obturation techniques. This may be attributed to additional cost involved or the lack of skill and training.
Discussion
The response rate was 72%. It was higher than in many previous surveys conducted in Western countries with better communication infrastructure [ 2 , 7 , 10 , 11 ].The vast majority of the respondents did not practice single visit root canal treatment. This finding was in agreement with the results of a previous study undertaken in another developing country, Sudan [ 9 ]. However, a study from the US [ 12 ] demonstrated a clear inclination to single visit endodontics, especially in cases without apical periodontitis. Single visit treatment appears to have gained more popularity and an increased credibility in the pre-clinical endodontic teaching in America and Europe [ 4 ]. Another survey [ 3 ], showed that a high percentage of Flemish dentists performed single visit root canal treatment. Multiple visit endodontic treatment could be a direct result of lacking adequate clinical time to complete the treatment in a single visit. The dentists may prefer to wait till the complete subsidence of pain and other symptoms before obturating the canal system. Another possible explanation could be that the initial visit was spent for treating the pain and acute symptoms [ 3 ].Although the application of rubber dam is always recommended as a standard during root canal treatment procedure to provide isolation, protection and improve visual access, only five dentists reported using rubber dam very occasionally and not as a routine practice. Similar findings were found in Sudan (2%) and among Flemish dentists (3.4%) [ 6 , 8 ]. However, 59% of American dentists [ 6 ], 60% of dentists in UK [ 13 ] and 57% of general dental practitioners in New Zealand [ 14 ] reported using rubber dam routinely in endodontic treatment. The reasons for not using rubber dam could be the extra cost, additional time, lack of adequate skills or training, absence of patient's acceptability or inadequate education in the undergraduate teaching curriculum. It was found that continuing education course attendees seem to be encouraged to use rubber dam [ 14 ].In the current survey, most general dental practitioners used hydrogen peroxide and sodium hypochlorite solutions as canal irrigants. The same result was demonstrated amongst dentists in Switzerland [ 11 ]. Sodium hypochlorite is recommended as the material of choice for irrigating the root canal system because of its effective antimicrobial and tissue solving action [ 15 ]. The selection of irrigant could be associated with the use of rubber dam, as it was found that 70% of rubber dam users among British dentists irrigated with sodium hypochlorite, whilst non-users tended to use local anesthetic solution [ 13 ]. The current findings do not mirror these findings. The vast majority of our respondents were non-users of rubber dam and one third of them use sodium hypochlorite routinely. A similar trend toward using sodium hypochlorite as an irrigant despite not using rubber dam for isolation, was noticed amongst Flemish dentists [ 16 ]. In the UK, the majority of dentists used local anesthetic solution to irrigate the canal space [ 2 ].The use of either sodium hypochlorite or hydrogen peroxide without isolating the field of operation tightly with a rubber dam presents an obviously hazardous practice in the use of potentially irritant irrigation solutions.Despite the fact that calcium hydroxide is recognized as the standard intracanal medicament for inter-appointment dressing [ 17 ], it was used by only 11.5% of the respondents. More than one third of the general practitioners reported using formaldehyde-containing materials. This finding is consistent with previous findings recorded for Sudanese dentists [ 9 ]. Although formaldehyde-containing products have been used for their antimicrobial and fixative properties, they are toxic to periradicular tissues [ 18 ] and may have mutagenic and carcinogenic potential [ 19 ]. The use of calcium hydroxide, as intracanal medication, should be encouraged among dentists in developing countries such as Jordan, as it is effective against most root canal pathogens and able to denature bacterial endotoxins [ 20 , 21 ]. It has, also, been reported to be the material of choice by dentists in the Western world [ 11 , 22 ].The step back technique was the most popular canal preparation technique among North Jordanian general dental practitioners. The filing (push-pull) technique, on the other hand, was used by 27.5% of the respondents. In another study, 60.4% of Flemish dentists used the standard filing technique [ 16 ]. Generally, dentists in Jordan tended to use hand instruments and were not inclined to use more advanced engine driven techniques for shaping the root canal system.Almost half of the general dental practioners in North Jordan used cold lateral compaction of gutta-percha to obturate the root canal space. This technique is acknowledged universally and is the most common obturation technique [ 4 ]. However, 31.3% of the dentists in the current survey used a single cone technique, in common with 68% of Swiss dentists [ 11 ]. Additionally, 12.2% of respondents used only paste to obturate the root canal system. Seemingly, dentists in North Jordan are not strong advocates of the more recently introduced advanced obturation techniques. This may be attributed to additional cost involved or the lack of skill and training.Conclusions This study investigated the status of endodontic practice among general dental practitioners working in private offices in North Jordan. It demonstrated that dentists performed procedures which often deviated from well-acknowledged endodontic quality guidelines. Dentists did not use rubber dam for isolation and frequently use formaldehyde-containing materials for inter-appointments dressing. In addition, a significant proportion of dentists (n = 30) did not use radiographs at any stage of endodontic treatment. General practitioners did not seem to keep up with recently introduced techniques, but use more conventional methods. The North Jordanian general dental practitioners carried out endodontic treatment with few referals to specialists. However, the absence of postgraduate endodontic programs and continuing education courses in addition to economic restrictions could explain why dentists in Jordan do not carry out endodontic treatment in accordance with recognized international standards.
Conclusions
This study investigated the status of endodontic practice among general dental practitioners working in private offices in North Jordan. It demonstrated that dentists performed procedures which often deviated from well-acknowledged endodontic quality guidelines. Dentists did not use rubber dam for isolation and frequently use formaldehyde-containing materials for inter-appointments dressing. In addition, a significant proportion of dentists (n = 30) did not use radiographs at any stage of endodontic treatment. General practitioners did not seem to keep up with recently introduced techniques, but use more conventional methods.The North Jordanian general dental practitioners carried out endodontic treatment with few referals to specialists. However, the absence of postgraduate endodontic programs and continuing education courses in addition to economic restrictions could explain why dentists in Jordan do not carry out endodontic treatment in accordance with recognized international standards.Competing Interests None declared.
Competing Interests
None declared.Pre-publication history The pre-publication history for this paper can be accessed here:
Pre-publication history
The pre-publication history for this paper can be accessed here:Supplementary Material Additional File 1 Endodontic Survey Questionnaire text of the file contains questions related to endodontic practice among general dental practitioners. Click here for file
Supplementary Material
Additional File 1
Endodontic Survey Questionnaire text of the file contains questions related to endodontic practice among general dental practitioners.Click here for file


Thanks to the tools of molecular biology, our understanding of the 100-plus diseases known collectively as cancer has increased dramatically over the past decades. While each of these cancers exhibits unique characteristics reflecting the particular cell or tissue it springs from, the disease follows a similar arc in nearly all its forms. Cancer is a multistep disease that begins when genetic damage—initiated by a multitude of agents—unleashes a single cell from the normal constraints on cellular proliferation. This single transformed cell generates a colony of similarly abnormal progeny that can take decades to develop into malignancies.While events that stimulate uncontrolled cell division can promote cancer, mutations in tumor suppressor genes figure prominently in tumor progression. Disruptions in the pRb (retinoblastoma 1) tumor suppressor, for example, are often seen early in cancer development, sensitizing cells to tumorigenesis. pRb, along with other “pocket proteins”—so-called because they share an amino acid domain called the Rb pocket—regulate cell cycle progression, apoptosis (programmed cell death), and cellular differentiation. Some tumor suppressors, such as p53, can trigger apoptosis, ultimately sacrificing cells that have sustained DNA damage or other types of cellular stress.Mutations in both the pRb and p53 tumor suppressor pathways are commonly seen in human cancers, though their interactions appear to vary depending on the tissue. In mouse brain epithelial cells, for example, loss of p53 function coupled with loss of pRb results in reduced apoptosis and increased tumor growth, while p53 loss in mouse brain astrocytes (cells that support neurons) does not affect tumor growth. Building on this work, Terry Van Dyke and colleagues report that loss of the pRb tumor suppressor in mammary tissue has the same effect—predisposition to tumor formation—seen in these other cell types. Despite the different environment inherent in each cell type, the initial events following loss of the pRb pathway were the same: increased proliferation and apoptosis, followed by tumorigenesis. But, surprisingly, pRb and p53 interactions varied in different cell types.Like most cancers, mammary gland cancer has a long latency period, prompting the researchers to ask what events engineer tumor progression. To investigate the relative contribution of pRb and p53 in tumorigenesis, the researchers generated a novel mouse model with a dysfunctional pRb pathway and various levels of p53 function in several cell types. This is a significant achievement in itself, as many agents that inactivate the pRb pathway also disrupt the p53 pathway. pRb inactivation, they show, causes abnormalities in mammary cell proliferation, apoptosis, and tissue morphology. In these mammary-specific pRb-deficient mice, p53 was responsible for most of the apoptotic response—decreased levels of p53 resulted in reduced apoptosis and accelerated tumorigenesis, but had no effect on proliferation. Interestingly, in other mouse models where aberrant proliferation is caused by disabling other pathways, loss of p53 was associated with increased proliferation—rather than reduced apoptosis—and early tumor formation. And while p53 is the main effector of apoptosis in brain and mammary epithelial cells, this is not the case in all tissues: in astrocytes, for example, the tumor suppressor Pten regulates apoptosis in response to pRb inactivation. Together these results indicate that specific cellular responses to a cancer-causing stimulus vary depending on the nature of the initial genetic injury and the cell type and that pRb and p53 interact in different ways in different tissues. And p53, it appears, contributes to tumor suppression—and thus progression—through multiple mechanisms.By creating a mouse model that disentangles the pRb and p53 pathways, Van Dyke and colleagues have added a valuable resource for studying breast cancer. This model, they propose, will facilitate further investigations into the relative contributions of these overlapping pathways to cancer progression. What's more, the model offers a vehicle for examining how pRb interacts with other breast cancer mutations, like the inherited mutations in the human  BRCA1  and  BRCA2  genes, to shed light on the complex series of events that ultimately cause breast cancer.BRCA1BRCA2


The administration of granulocyte colony-stimulating factor (G-CSF) to peripheral blood progenitor cell (PBPC) donors causes spleen length to increase, but the duration of enlargement is not known. Eighteen healthy subjects were given 10 μg/kg of G-CSF for 5 days and a PBSC concentrate was collected by apheresis. Ultrasound scans were used to assess craniocaudal spleen length before and after G-CSF administration. Mean spleen length increased from a baseline length of 10.7 ± 1.3 cm to 12.1 ± 1.2 cm on the apheresis day (p < 0.001). Ten days after apheresis, spleen length fell to 10.5 ± 1.2 cm and did not differ from baseline levels (p = 0.57), but in 3 subjects remained 0.5 cm greater than baseline length. Increases in spleen length in PBPC donors are transient and reversible.

Background Peripheral blood progenitor cell (PBPC) concentrates donors are routinely given granulocyte colony-stimulating factor (G-CSF) to increase the concentration of circulating PBPCs and hence the number of progenitors that can be collected by apheresis. Typically 10 to 16 μg/kg of G-CSF are given subcutaneously daily for 4 to 6 days prior to the collection [ 1 - 3 ]. The administration of G-CSF to healthy PBPC concentrates donors is very safe, but there have been four reports of spontaneous rupture of the spleen and splenectomy in healthy allogeneic PBPC donors given G-CSF [ 4 - 7 ]. While spontaneous rupture of the spleen in PBSC donors given G-CSF is rare, the administration of G-CSF for five days causes spleen length to increase in almost all healthy donors [ 8 , 9 ]. The increase in length is highly variable, but the mean increase is approximately 13%. Spleen length begins to return to baseline levels quickly, but it is not known how long it takes to return to baseline. In a previous study of 20 PBPC donors given 10 μg/kg of G-CSF for 5 days, we found that spleen length measured four days after the last dose of G-CSF was less than the length on the day of apheresis but greater than baseline values [ 8 ]. Since allogeneic PBPC donors may be at risk for splenic rupture while the spleen is enlarged, it is important to determine when spleen size returns to baseline levels. The purpose of this study was to determine if spleen length returns to baseline 10 days after G-CSF-mobilized PBPC concentrates are collected by apheresis from healthy subjects.
Background
Peripheral blood progenitor cell (PBPC) concentrates donors are routinely given granulocyte colony-stimulating factor (G-CSF) to increase the concentration of circulating PBPCs and hence the number of progenitors that can be collected by apheresis. Typically 10 to 16 μg/kg of G-CSF are given subcutaneously daily for 4 to 6 days prior to the collection [ 1 - 3 ]. The administration of G-CSF to healthy PBPC concentrates donors is very safe, but there have been four reports of spontaneous rupture of the spleen and splenectomy in healthy allogeneic PBPC donors given G-CSF [ 4 - 7 ].While spontaneous rupture of the spleen in PBSC donors given G-CSF is rare, the administration of G-CSF for five days causes spleen length to increase in almost all healthy donors [ 8 , 9 ]. The increase in length is highly variable, but the mean increase is approximately 13%. Spleen length begins to return to baseline levels quickly, but it is not known how long it takes to return to baseline. In a previous study of 20 PBPC donors given 10 μg/kg of G-CSF for 5 days, we found that spleen length measured four days after the last dose of G-CSF was less than the length on the day of apheresis but greater than baseline values [ 8 ].Since allogeneic PBPC donors may be at risk for splenic rupture while the spleen is enlarged, it is important to determine when spleen size returns to baseline levels. The purpose of this study was to determine if spleen length returns to baseline 10 days after G-CSF-mobilized PBPC concentrates are collected by apheresis from healthy subjects.Methods Study design All of the subjects were in good health and were donating G-CSF-mobilized PBPC concentrates for laboratory investigations. The donors were given 10 μg/kg of G-CSF (Filgrastim, Amgen, Thousand Oaks, CA) daily for 5 days, and a PBSC concentrate was collected approximately 2 hours after the last G-CSF dose was given. PBPC concentrates were collected with a CS3000 blood cell separator (Baxter Health Care Corporation, Round Lake, IL). Spleen length was evaluated by ultrasound examination three times: prior to the administration of the first dose of G-CSF, on the day of apheresis, and 10 or 11 days after apheresis. This study was approved by the Institutional Review Board of the National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland. Spleen length assessment Craniocaudal spleen length was assessed using ultrasound (Acuson Aspen Advanced, Siemens Medical Solutions Ultrasound Division, Mountain View, CA) with a sector transducer (Acuson, 4V1, 4.0 mHz frequency, Siemens Medical Solutions). The intra-observer error for measuring spleen length using ultrasound is 4.9 mm when healthy subjects are evaluated at separate settings [ 10 ]. Blood counts and chemistries Complete blood counts were performed with an automated cell counter (Cell Dyne 4000, Abbott Diagnostics, Santa Clara, CA). CD34+ cell counts were performed using a flow cytometer (Beckman Coulter, Miami, FL). Statistical analysis Spleen lengths measured before and after the G-CSF course were compared using 2-tailed paired t-tests. Spleen length changes were also compared among males and females and Caucasians and non-Caucasians using 2-tailed t-tests. The percent change in spleen length was compared with blood counts, CD34+ cell counts, and donor age using linear regression.
Methods
Study design All of the subjects were in good health and were donating G-CSF-mobilized PBPC concentrates for laboratory investigations. The donors were given 10 μg/kg of G-CSF (Filgrastim, Amgen, Thousand Oaks, CA) daily for 5 days, and a PBSC concentrate was collected approximately 2 hours after the last G-CSF dose was given. PBPC concentrates were collected with a CS3000 blood cell separator (Baxter Health Care Corporation, Round Lake, IL). Spleen length was evaluated by ultrasound examination three times: prior to the administration of the first dose of G-CSF, on the day of apheresis, and 10 or 11 days after apheresis. This study was approved by the Institutional Review Board of the National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.
Study design
All of the subjects were in good health and were donating G-CSF-mobilized PBPC concentrates for laboratory investigations. The donors were given 10 μg/kg of G-CSF (Filgrastim, Amgen, Thousand Oaks, CA) daily for 5 days, and a PBSC concentrate was collected approximately 2 hours after the last G-CSF dose was given. PBPC concentrates were collected with a CS3000 blood cell separator (Baxter Health Care Corporation, Round Lake, IL). Spleen length was evaluated by ultrasound examination three times: prior to the administration of the first dose of G-CSF, on the day of apheresis, and 10 or 11 days after apheresis. This study was approved by the Institutional Review Board of the National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.Spleen length assessment Craniocaudal spleen length was assessed using ultrasound (Acuson Aspen Advanced, Siemens Medical Solutions Ultrasound Division, Mountain View, CA) with a sector transducer (Acuson, 4V1, 4.0 mHz frequency, Siemens Medical Solutions). The intra-observer error for measuring spleen length using ultrasound is 4.9 mm when healthy subjects are evaluated at separate settings [ 10 ].
Spleen length assessment
Craniocaudal spleen length was assessed using ultrasound (Acuson Aspen Advanced, Siemens Medical Solutions Ultrasound Division, Mountain View, CA) with a sector transducer (Acuson, 4V1, 4.0 mHz frequency, Siemens Medical Solutions). The intra-observer error for measuring spleen length using ultrasound is 4.9 mm when healthy subjects are evaluated at separate settings [ 10 ].Blood counts and chemistries Complete blood counts were performed with an automated cell counter (Cell Dyne 4000, Abbott Diagnostics, Santa Clara, CA). CD34+ cell counts were performed using a flow cytometer (Beckman Coulter, Miami, FL).
Blood counts and chemistries
Complete blood counts were performed with an automated cell counter (Cell Dyne 4000, Abbott Diagnostics, Santa Clara, CA). CD34+ cell counts were performed using a flow cytometer (Beckman Coulter, Miami, FL).Statistical analysis Spleen lengths measured before and after the G-CSF course were compared using 2-tailed paired t-tests. Spleen length changes were also compared among males and females and Caucasians and non-Caucasians using 2-tailed t-tests. The percent change in spleen length was compared with blood counts, CD34+ cell counts, and donor age using linear regression.
Statistical analysis
Spleen lengths measured before and after the G-CSF course were compared using 2-tailed paired t-tests. Spleen length changes were also compared among males and females and Caucasians and non-Caucasians using 2-tailed t-tests. The percent change in spleen length was compared with blood counts, CD34+ cell counts, and donor age using linear regression.Results and Discussion The median age of the 18 healthy subjects was 34 years old and ranged from 22 to 55 years of age. Eight of the subjects were male, 13 of the donors were Caucasian, 3 were African American, and 2 Asian. Apheresis day spleen length increased above baseline length in 17 of 18 donors (Figure  1 ). The mean spleen length increased from a baseline level of 10.7 ± 1.3 cm to 12.1 ± 1.2 cm on the day of apheresis (p < 0.001). The mean increase in length was 1.4 ± 0.8 cm or 13.1 ± 8.9%. Figure 1 Spleen length changes in healthy subjects donating G-CSF-mobilized PBPC concentrates. Eighteen subjects were given 10 μg/kg of G-CSF for 5 days and a PBPC concentrate was collected approximately 2 hours after the last dose of G-CSF. Spleen length was measured by ultrasound before G-CSF was given (day 1), immediately after the PBPC concentrate collection (day 5), and approximately 10 or 11 days after the collection (day 15 or 16). Spleen length was measured 10 or 11 days after apheresis in all 18 subjects. The mean spleen length 10 days after apheresis fell to 10.5 ± 1.2 cm and was less than the apheresis day spleen length (p < 0.001). There was no difference between the 10-day post-apheresis and pre-G-CSF spleen length (p = 0.57). The spleen length 10 days after apheresis was less than the apheresis day length in all 17 donors whose spleen length increased. However, the spleen length 10 days after apheresis remained more than 0.5 cm greater than baseline spleen length in 3 subjects (Table  1 ). All 3 were Caucasian and 2 were female. Their spleen length remained 6.7%, 11.0%, and 10.5% greater than baseline levels. Two of these subjects had relatively large increases in spleen length of 18.3% and 26.3%, but their spleen length fell considerably 10 days after apheresis. It is likely that the spleen returned to baseline length in these 2 subjects shortly after the third ultrasound was preformed. The other subject's spleen increased only 12.0% in length and 10 days after apheresis had changed little. It is not certain when this subject's spleen returned to baseline length. Table 1 Peripheral Blood Stem Cell Donors Whose Spleen Length 10 Days After Apheresis Remained More than 0.5 cm Greater than Baseline Length Spleen Length (cm) Donor Age (Yrs) Gender Race Baseline Apheresis Day 10 Days Post-Apheresis Enlargement 10 Days Post-Apheresis 7 23 Female Cauc 10.4 12.3 11.1 0.7 13 22 Female Cauc 10.0 11.2 11.1 1.1 14 54 Male Cauc 9.5 12.0 10.5 1.0 Cauc = Caucasian PBPC donors with the largest increase in spleen size may be at the greatest risk for spontaneous splenic rupture. In order to determine if subject age, gender, race, or post-G-CSF blood counts affected the magnitude of spleen enlargement, we assessed the relationship between these factors and percent change in spleen length in the 18 subjects in this study and 20 subjects studied previously [14]. There was no difference in the spleen length increase between males and females (12.3 ± 9.7% versus 14.8 ± 8.0%, p = 0.40) or between Caucasians and non-Caucasians (13.5 ± 8.4% versus 13.6 ± 10.4%; p = 0.98). Spleen length increase was not related to donor age (r = 0.13). In addition, spleen length increase was not related to preapheresis CD34+ (r = 0.04), WBC (r = 0.05), neutrophil (r = 0.07), lymphocyte (r = -0.14), monocyte (r = -0.04), and platelet counts (r = 0.19) or hemoglobin level (r = -0.04).
Results and Discussion
The median age of the 18 healthy subjects was 34 years old and ranged from 22 to 55 years of age. Eight of the subjects were male, 13 of the donors were Caucasian, 3 were African American, and 2 Asian. Apheresis day spleen length increased above baseline length in 17 of 18 donors (Figure  1 ). The mean spleen length increased from a baseline level of 10.7 ± 1.3 cm to 12.1 ± 1.2 cm on the day of apheresis (p < 0.001). The mean increase in length was 1.4 ± 0.8 cm or 13.1 ± 8.9%.Spleen length was measured 10 or 11 days after apheresis in all 18 subjects. The mean spleen length 10 days after apheresis fell to 10.5 ± 1.2 cm and was less than the apheresis day spleen length (p < 0.001). There was no difference between the 10-day post-apheresis and pre-G-CSF spleen length (p = 0.57). The spleen length 10 days after apheresis was less than the apheresis day length in all 17 donors whose spleen length increased. However, the spleen length 10 days after apheresis remained more than 0.5 cm greater than baseline spleen length in 3 subjects (Table  1 ). All 3 were Caucasian and 2 were female. Their spleen length remained 6.7%, 11.0%, and 10.5% greater than baseline levels. Two of these subjects had relatively large increases in spleen length of 18.3% and 26.3%, but their spleen length fell considerably 10 days after apheresis. It is likely that the spleen returned to baseline length in these 2 subjects shortly after the third ultrasound was preformed. The other subject's spleen increased only 12.0% in length and 10 days after apheresis had changed little. It is not certain when this subject's spleen returned to baseline length.Peripheral Blood Stem Cell Donors Whose Spleen Length 10 Days After Apheresis Remained More than 0.5 cm Greater than Baseline LengthDonorAge (Yrs)GenderRaceBaselineApheresis Day10 Days Post-ApheresisEnlargement 10 Days Post-ApheresisCauc = CaucasianPBPC donors with the largest increase in spleen size may be at the greatest risk for spontaneous splenic rupture. In order to determine if subject age, gender, race, or post-G-CSF blood counts affected the magnitude of spleen enlargement, we assessed the relationship between these factors and percent change in spleen length in the 18 subjects in this study and 20 subjects studied previously [14]. There was no difference in the spleen length increase between males and females (12.3 ± 9.7% versus 14.8 ± 8.0%, p = 0.40) or between Caucasians and non-Caucasians (13.5 ± 8.4% versus 13.6 ± 10.4%; p = 0.98). Spleen length increase was not related to donor age (r = 0.13). In addition, spleen length increase was not related to preapheresis CD34+ (r = 0.04), WBC (r = 0.05), neutrophil (r = 0.07), lymphocyte (r = -0.14), monocyte (r = -0.04), and platelet counts (r = 0.19) or hemoglobin level (r = -0.04).Conclusions Healthy PBPC concentrate donors given G-CSF should be warned that their spleens will be enlarged for a brief time and that they may be at risk of splenic rupture. Most donors are likely at risk for splenic rupture only during the time of G-CSF administration and for about 10 days after the completion of the G-CSF course. Since splenic enlargement may persist for longer periods in some donor, until more data are available it may be worthwhile to counsel PBPC donors to avoid activities that could lead to abdominal and splenic trauma for 2 to 3 weeks after the last dose of G-CSF.
Conclusions
Healthy PBPC concentrate donors given G-CSF should be warned that their spleens will be enlarged for a brief time and that they may be at risk of splenic rupture. Most donors are likely at risk for splenic rupture only during the time of G-CSF administration and for about 10 days after the completion of the G-CSF course. Since splenic enlargement may persist for longer periods in some donor, until more data are available it may be worthwhile to counsel PBPC donors to avoid activities that could lead to abdominal and splenic trauma for 2 to 3 weeks after the last dose of G-CSF.


One of the most common and most serious complications of both type 1 and type 2 diabetes is diabetic nephropathy. It occurs in around 30% of patients with type 1 diabetes and 10% to 40% of patients with type 2 diabetes. Diabetic nephropathy is the leading cause of renal failure in the developed world. The main effect of diabetic nephropathy is proteinuria, initially in very small amounts but which increases, leading to nephrotic syndrome and end-stage renal disease in most cases.Various risk factors in individuals with diabetes are known to increase the chance of developing diabetic nephropathy, including South Asian or African background, male sex, long history of diabetes, poor blood sugar control, high blood pressure, and smoking. One early change associated with diabetic nephropathy is degeneration of the renal tubular epithelium, but the exact cause of this at the cellular level is unclear. Erwin Böttinger and colleagues have dissected out one key point in the progression to diabetic nephropathy. They looked at cell lines of renal tubular cells from humans and mice and kidney biopsies from patients with diabetic nephropathy, patients with non-diabetic renal disease, and mice with genetic and induced diabetes. In the human cell lines they showed that glucose induced the expression of CD36, a receptor known to have a role in adhesion and signal transduction (in addition to being the receptor for malaria-infected erythrocytes). They then went on to show that apoptosis of these cells occurred in the presence of glycated (glucose-modified) albumins or free fatty acids, which are present in increased amounts in patients with diabetes, and that CD36 was essential for the apoptosis to occur. They then examined how CD36 triggered apoptosis and found that it involved src kinase, p38 MAP kinase, and caspase 3.Comparing mice and humans, the researchers found that the two species are not alike: diabetic mice did not show an increase in tubular expression of CD36—even though the gene is present in mice—and had normal tubular epithelium and no tubular apoptosis. They confirmed this difference between humans and mice by showing that normal mouse epithelial cell lines were resistant to apoptosis caused by the glycated albumins; however, artificially expressing CD36 in these lines made them susceptible to apoptosis by these modified albumins.These results provide insight into one of the crucial steps in diabetic nephropathy and, in humans at least, might help to explain why high blood glucose is so damaging to the kidney, hence providing a good reason—if another is needed—for encouraging patients to control blood glucose as tightly as possible.


The Drugs for Neglected Diseases Initiative is a new, public- sector organization dedicated to drug discovery

In wealthy countries, state-funded research has yielded breakthroughs in molecular biology, chemistry, and engineering. These advances have been taken up by the pharmaceutical industry and applied to drug development for a growing range of illnesses and conditions. As a result, patients have access to new drugs that are better tolerated, more specific, and more effective than old ones.In poor countries, however, millions of people have yet to experience the benefits wrought by science. The deadly infectious diseases that plague them, such as sleeping sickness, Chagas disease, and visceral leishmaniasis, fail to arouse the interest of drug developers.The Drugs for Neglected Diseases Initiative (DNDi) is a new, not-for-profit organisation set up to correct this fatal imbalance by developing new drugs for these forgotten patients.Dropped off the Radar Screen Most of the drugs still used to treat ‘neglected diseases’ were developed in colonial times. These are often expensive, difficult to administer, and hard to tolerate; several of them are also becoming ineffective because of increasing parasite resistance. Very few new alternatives have been developed in the past decades: between 1975 and 1999, 1,393 new drugs were made available to the public, but only 16 of these were meant for neglected diseases [ 1 ]. What makes the lack of drugs more difficult to accept is that scientists know an enormous amount about kinetoplastids, the organisms responsible for sleeping sickness, Chagas disease, and leishmaniasis [ 2 ]. The wealth of knowledge generated in this field could easily be used for drug development if the treatment of neglected diseases were perceived as financially attractive. But populations affected by neglected diseases have no purchasing power, so there is no financial incentive for drug companies to develop the drugs. The basic mechanics of the market-driven system are failing to help these populations. So most scientific research stops at the publication stage or falls through the gaps at different stages of the drug development pipeline ( Figure 1 ) [ 3 ]. Figure 1 The Drug Development Pipeline Because of the gaps in the development pipeline, potential new drugs for neglected diseases often stay stuck at an early stage of development. (Photos: World Health Organization/P.Virot and World Health Organization/Eric Miller)
Dropped off the Radar Screen
Most of the drugs still used to treat ‘neglected diseases’ were developed in colonial times. These are often expensive, difficult to administer, and hard to tolerate; several of them are also becoming ineffective because of increasing parasite resistance. Very few new alternatives have been developed in the past decades: between 1975 and 1999, 1,393 new drugs were made available to the public, but only 16 of these were meant for neglected diseases [ 1 ].What makes the lack of drugs more difficult to accept is that scientists know an enormous amount about kinetoplastids, the organisms responsible for sleeping sickness, Chagas disease, and leishmaniasis [ 2 ]. The wealth of knowledge generated in this field could easily be used for drug development if the treatment of neglected diseases were perceived as financially attractive. But populations affected by neglected diseases have no purchasing power, so there is no financial incentive for drug companies to develop the drugs. The basic mechanics of the market-driven system are failing to help these populations. So most scientific research stops at the publication stage or falls through the gaps at different stages of the drug development pipeline ( Figure 1 ) [ 3 ].Whose Role Is It, Anyway? It is dangerous to oversimplify the causes of this situation. What share of responsibility for the world's health is borne by the pharmaceutical industry, which has the know-how and the resources for innovation? Aren't international organisations also partly responsible? After all, they are the ones who allocate major funding for health programmes and encourage research programmes. And what about public research institutions in rich countries that generate the knowledge used by industry? Governments have the power to influence their research priorities and drug development decisions, either through funding or direct involvement. Unfortunately drugs for neglected diseases are low priority for governments [ 4 ]. They tend to prioritise research with potential commercial applications instead.
Whose Role Is It, Anyway?
It is dangerous to oversimplify the causes of this situation. What share of responsibility for the world's health is borne by the pharmaceutical industry, which has the know-how and the resources for innovation? Aren't international organisations also partly responsible? After all, they are the ones who allocate major funding for health programmes and encourage research programmes. And what about public research institutions in rich countries that generate the knowledge used by industry? Governments have the power to influence their research priorities and drug development decisions, either through funding or direct involvement. Unfortunately drugs for neglected diseases are low priority for governments [ 4 ]. They tend to prioritise research with potential commercial applications instead.Responses to the Crisis All is not doom and gloom. In the past few years, there has been some movement on research and development (R&D) for neglected diseases. Despite its broad mandate and limited resources, the Special Programme for Research and Training in Tropical Diseases (TDR)—established and funded by the World Health Organization, the World Bank, and the United Nations Development Programme—can be credited with several important successes in the fight against malaria and leishmaniasis. The Medicines for Malaria Venture and the Global Alliance for TB Drug Development were set up as public–private partnerships to tackle malaria and tuberculosis. These partnerships were made possible by the fact that malaria and tuberculosis are global diseases, affecting patients in the North and South, so there was enough of a market to persuade industry to develop new drugs for these diseases. A different solution, however, was needed for diseases that are limited to tropical countries, are of no military or strategic interest to wealthy countries, and are not supported by markets or patients' organisations capable of attracting the attention of politicians. This is the kind of solution put forward by the DNDi.
Responses to the Crisis
All is not doom and gloom. In the past few years, there has been some movement on research and development (R&D) for neglected diseases. Despite its broad mandate and limited resources, the Special Programme for Research and Training in Tropical Diseases (TDR)—established and funded by the World Health Organization, the World Bank, and the United Nations Development Programme—can be credited with several important successes in the fight against malaria and leishmaniasis. The Medicines for Malaria Venture and the Global Alliance for TB Drug Development were set up as public–private partnerships to tackle malaria and tuberculosis. These partnerships were made possible by the fact that malaria and tuberculosis are global diseases, affecting patients in the North and South, so there was enough of a market to persuade industry to develop new drugs for these diseases.A different solution, however, was needed for diseases that are limited to tropical countries, are of no military or strategic interest to wealthy countries, and are not supported by markets or patients' organisations capable of attracting the attention of politicians. This is the kind of solution put forward by the DNDi.A Collaborative Not-for-Profit DNDi is a not-for-profit organisation designed to mobilise resources for R&D of new drugs for neglected diseases. Many people and organisations around the world share an ambition to redress the lack of new treatments for neglected diseases, and bring the benefits of science to forgotten patients. Several of them came together to create DNDi: one humanitarian organisation—Médecins Sans Frontières; five research institutions—the Oswaldo Cruz Foundation from Brazil, the Indian Council for Medical Research, the Kenya Medical Research Institute, the Ministry of Health Malaysia, and the Pasteur Institute from France; and the TDR ( Box 1 ). Box 1. From Pipeline to Patients—Some Key Organizations
 DNDi:
 http://www.dndi.org

 TDR:
 http://www.who.int/tdr

 Medicines for Malaria Venture:
 http://www.mmv.org

 Global Alliance for TB Drug Development:
 http://www.tballiance.org

 Oswaldo Cruz Foundation:
 http://www.fiocruz.br

 Indian Council for Medical Research:
 http://icmr.nic.in/home.htm

 Kenya Medical Research Institute:
 http://www.kemri.org

 Ministry of Health Malaysia:
 http://dph.gov.my/

 Pasteur Institute:
 http://www.pasteur.fr/externe

 Médecins Sans Frontières:
 http://www.msf.org
 The initiative is a virtual organisation with a growing network of academic and R&D expertise at its disposal. The different players involved in DNDi are bringing their knowhow in parasitology and clinical trials, their experience treating neglected patients, and their drug manufacturing capacity. They are pooling these resources to move drugs stuck in the pipeline all the way to the patients themselves. Pharmaceutical companies have a particularly important role to play: they possess vast repositories of molecules, the means to move from development to industrial production, and highly specialised teams of researchers. Their contribution will be crucial to the success of DNDi.
A Collaborative Not-for-Profit
DNDi is a not-for-profit organisation designed to mobilise resources for R&D of new drugs for neglected diseases.Many people and organisations around the world share an ambition to redress the lack of new treatments for neglected diseases, and bring the benefits of science to forgotten patients. Several of them came together to create DNDi: one humanitarian organisation—Médecins Sans Frontières; five research institutions—the Oswaldo Cruz Foundation from Brazil, the Indian Council for Medical Research, the Kenya Medical Research Institute, the Ministry of Health Malaysia, and the Pasteur Institute from France; and the TDR ( Box 1 ).Box 1. From Pipeline to Patients—Some Key Organizations
DNDi:
 http://www.dndi.orgDNDi:TDR:
 http://www.who.int/tdrTDR:Medicines for Malaria Venture:
 http://www.mmv.orgMedicines for Malaria Venture:Global Alliance for TB Drug Development:
 http://www.tballiance.orgGlobal Alliance for TB Drug Development:Oswaldo Cruz Foundation:
 http://www.fiocruz.brOswaldo Cruz Foundation:Indian Council for Medical Research:
 http://icmr.nic.in/home.htmIndian Council for Medical Research:Kenya Medical Research Institute:
 http://www.kemri.orgKenya Medical Research Institute:Ministry of Health Malaysia:
 http://dph.gov.my/Ministry of Health Malaysia:Pasteur Institute:
 http://www.pasteur.fr/externePasteur Institute:Médecins Sans Frontières:
 http://www.msf.orgMédecins Sans Frontières:The initiative is a virtual organisation with a growing network of academic and R&D expertise at its disposal. The different players involved in DNDi are bringing their knowhow in parasitology and clinical trials, their experience treating neglected patients, and their drug manufacturing capacity. They are pooling these resources to move drugs stuck in the pipeline all the way to the patients themselves. Pharmaceutical companies have a particularly important role to play: they possess vast repositories of molecules, the means to move from development to industrial production, and highly specialised teams of researchers. Their contribution will be crucial to the success of DNDi.Matching Needs and Opportunities DNDi is a needs-driven initiative—in other words, the needs of patients suffering from neglected diseases are paramount in its search for new drugs to treat them. The organisations that make up DNDi have firsthand knowledge of these needs because they work with patients in disease-endemic countries throughout the developing world. The initiative is taking this knowledge of patient needs, matching it with opportunities in R&D, and pushing the most relevant projects through the pipeline. Ultimately, neglected patients will have access to drugs targeting their specific diseases, drugs that were designed with them specifically in mind—such as short-course, low-toxicity treatments that don't require hospitalisation, or tablets to swallow rather than injections. To identify opportunities in R&D that are both relevant to patient needs and that meet required criteria of scientific merit, DNDi is sending out calls for letters of interest to the scientific community via advertisements in journals and posted on the DNDi website ( http://www.dndi.org ). These have already pinpointed several promising projects. DNDi is also proactively contacting scientists working on infectious diseases, and surveying published literature for research of interest.
Matching Needs and Opportunities
DNDi is a needs-driven initiative—in other words, the needs of patients suffering from neglected diseases are paramount in its search for new drugs to treat them. The organisations that make up DNDi have firsthand knowledge of these needs because they work with patients in disease-endemic countries throughout the developing world. The initiative is taking this knowledge of patient needs, matching it with opportunities in R&D, and pushing the most relevant projects through the pipeline. Ultimately, neglected patients will have access to drugs targeting their specific diseases, drugs that were designed with them specifically in mind—such as short-course, low-toxicity treatments that don't require hospitalisation, or tablets to swallow rather than injections.To identify opportunities in R&D that are both relevant to patient needs and that meet required criteria of scientific merit, DNDi is sending out calls for letters of interest to the scientific community via advertisements in journals and posted on the DNDi website ( http://www.dndi.org ). These have already pinpointed several promising projects. DNDi is also proactively contacting scientists working on infectious diseases, and surveying published literature for research of interest.In the Pipeline DNDi's project portfolio currently holds nine projects at different stages of development to address identified needs for the treatment of visceral leishmaniasis, sleeping sickness ( Box 2 ), Chagas disease, and malaria ( Figure 2 ). At discovery stage, DNDi is working on validating the kinetoplastid enzyme dihydrofolate reductase as a potential target for leishmaniasis, trypanosomiasis, and Chagas disease, and on identifying inhibitors of the kinetoplastid enzymes trypanothione reductase and protein farnesyltransferase. It is also conducting high throughput screening on whole cell trypanosomes to discover novel lead compounds. Box 2. New Drugs for Sleeping Sickness Only a few drugs exist to treat sleeping sickness, and they are toxic or difficult to administer. Melarsoprol kills one in 20 patients. Eflornithine requires four daily infusions over 14 days. Given these limited options, DNDi is focusing on identifying new compounds that can cross the blood–brain barrier to treat second stage sleeping sickness. DNDi is using high throughput screening on whole cell trypanosomes to discover novel lead compounds, and is working to identify and optimise inhibitors of the enzyme protein farnesyltransferase. The initiative is working on validating the kinetoplastid enzyme dihydrofolate reductase as a drug target. Identifying trypanothione inhibitors is also relevant to other trypanosome parasites. These are long term projects. Nifurtimox, a drug for Chagas disease, has been used to treat sleeping sickness since the 1970s in some isolated places. It has never been extended to more people because no one has studied its safety or effectiveness. DNDi will assess its short-term usefulness by conducting clinical trials on a treatment combination of eflornithine and nifurtimox. DNDi will continue to explore other short- and medium-term projects. Figure 2 DNDi Projects DNDi's project portfolio contains nine projects spread out across the drug development pipeline for the treatment of leishmaniasis, sleeping sickness, Chagas disease, and malaria. HAT, human African trypanosomiasis (sleeping sickness); VL, visceral leishmaniasis. The R&D of new drugs is time-consuming and expensive if the process starts at the early discovery stage, because of the associated risk of project attrition along the way. DNDi is therefore investing resources in several pre-development and development projects as well. These include developing fixed dose combinations of artesunate/amodiaquine and artesunate/mefloquine for use against chloroquine-resistant malaria in Africa and Asia, respectively; pushing for registration of paromomycin for use against visceral leishmaniasis in Africa; assessing combinations of existing drugs for visceral leishmaniasis; and evaluating the usefulness of nifurtimox in combination with eflornithine in the treatment of sleeping sickness.
In the Pipeline
DNDi's project portfolio currently holds nine projects at different stages of development to address identified needs for the treatment of visceral leishmaniasis, sleeping sickness ( Box 2 ), Chagas disease, and malaria ( Figure 2 ). At discovery stage, DNDi is working on validating the kinetoplastid enzyme dihydrofolate reductase as a potential target for leishmaniasis, trypanosomiasis, and Chagas disease, and on identifying inhibitors of the kinetoplastid enzymes trypanothione reductase and protein farnesyltransferase. It is also conducting high throughput screening on whole cell trypanosomes to discover novel lead compounds.Box 2. New Drugs for Sleeping Sickness
Only a few drugs exist to treat sleeping sickness, and they are toxic or difficult to administer. Melarsoprol kills one in 20 patients. Eflornithine requires four daily infusions over 14 days. Given these limited options, DNDi is focusing on identifying new compounds that can cross the blood–brain barrier to treat second stage sleeping sickness.DNDi is using high throughput screening on whole cell trypanosomes to discover novel lead compounds, and is working to identify and optimise inhibitors of the enzyme protein farnesyltransferase. The initiative is working on validating the kinetoplastid enzyme dihydrofolate reductase as a drug target. Identifying trypanothione inhibitors is also relevant to other trypanosome parasites. These are long term projects.Nifurtimox, a drug for Chagas disease, has been used to treat sleeping sickness since the 1970s in some isolated places. It has never been extended to more people because no one has studied its safety or effectiveness. DNDi will assess its short-term usefulness by conducting clinical trials on a treatment combination of eflornithine and nifurtimox. DNDi will continue to explore other short- and medium-term projects.The R&D of new drugs is time-consuming and expensive if the process starts at the early discovery stage, because of the associated risk of project attrition along the way. DNDi is therefore investing resources in several pre-development and development projects as well. These include developing fixed dose combinations of artesunate/amodiaquine and artesunate/mefloquine for use against chloroquine-resistant malaria in Africa and Asia, respectively; pushing for registration of paromomycin for use against visceral leishmaniasis in Africa; assessing combinations of existing drugs for visceral leishmaniasis; and evaluating the usefulness of nifurtimox in combination with eflornithine in the treatment of sleeping sickness.Advocacy for Change Governments can—some might say should—influence drug development choices. DNDi strongly believes that governments in both developed and developing countries should take an active interest in the R&D of new drugs for neglected diseases. In parallel to its own drug development activities, DNDi is working to raise awareness of the neglected disease crisis among key policy- and decision-makers, for instance the European Commission and the National Institutes for Health in the United States.
Advocacy for Change
Governments can—some might say should—influence drug development choices. DNDi strongly believes that governments in both developed and developing countries should take an active interest in the R&D of new drugs for neglected diseases. In parallel to its own drug development activities, DNDi is working to raise awareness of the neglected disease crisis among key policy- and decision-makers, for instance the European Commission and the National Institutes for Health in the United States.Conclusion In the poorer countries in the world, over 350 million people are at risk from neglected diseases. Currently available treatments are inadequate or nonexistent, and new solutions are urgently needed. DNDi is working to ensure that the advances of science that have brought health and comfort to wealthy nations also benefit these neglected populations.
Conclusion
In the poorer countries in the world, over 350 million people are at risk from neglected diseases. Currently available treatments are inadequate or nonexistent, and new solutions are urgently needed. DNDi is working to ensure that the advances of science that have brought health and comfort to wealthy nations also benefit these neglected populations.