In this study, we used fl/fl;Alb-Cre+/−Met livers. Up-regulation of stress response  and early growth response  genes as assessed by RT-qPCR and/or microarray profiling was unchanged. This was consistent with an early induction of MAPK/Erk and STAT3. However, after a successful completion of the first round of DNA replication, c-Met deficient hepatocytes were blocked in early/mid G2 phase as shown by staining with phosphorylated form of histone H3. Furthermore, loss of c-Met in hepatocytes diminished the subsequent G1/S progression and delayed liver recovery after partial hepatectomy. Upstream signaling pathways involved in the blockage of G2/M transition included lack of persistent Erk1/2 activation and inability to up-regulate the levels of Cdk1, Plk1, Aurora A and B, and Mad2 along with a defective histone 3 phosphorylation and lack of chromatin condensation. Continuous supplementation with EGF in vitro increased proliferation of fl/fl;Alb-Cre+/−Met primary hepatocytes and partially restored expression levels of mitotic cell cycle regulators albeit to a lesser degree as compared to control cultures.The priming events appeared to be intact in In conclusion, our results assign a novel non-redundant function for HGF/c-Met signaling in regulation of G2/M gene expression program via maintaining a persistent Erk1/2 activation throughout liver regeneration. Among the major growth factors involved at this stage is the hepatocyte growth factor (HGF) Liver regeneration after partial hepatectomy (PH) is a process of compensatory hyperplasia which involves all cell types within the remaining liver lobes and depends on the multitude of interrelated regulatory pathways that directly or indirectly control successful restoration of hepatic mass HGF is a mesenchyme-derived multifunctional molecule that elicits its activities through a single receptor c-Met c-met using a ubiquitously expressed alpha/beta interferon-inducible promoter resulted in suppressed Erk1/2 activation and defective exit from quiescence in regenerating mouse liver c-met via RNA interference c-met in diverse liver cell types, and therefore the extent to which the lack of c-Met signaling specifically in hepatocytes contributed to impaired cell cycle regulation could not be elucidated. Further, although prior evidence implicated Erk1/2 pathway in regulating G1/S transition, its role during G2/M transition and how it relates to c-Met function remains to be fully realized.A key step in c-Met signaling is HGF-induced phosphorylation of the kinase domain followed by autophosphorylation and activation of numerous downstream effectors c-met was deleted either by a constitutive Alb-Cre or inducible Mx1-Cre transgenes. Here we report for the first time that besides its role in G1/S progression, c-Met is essential regulator of G2/M progression through a mechanism involving Erk1/2-dependent activation of key mitotic regulators including Cdk1, Plk1, and Aurora A and B.To address these questions, we designed our experiments using two c-Met conditional knockout models: a hepatocyte- and liver specific mutant mice in which the exon encoding the ATP-binding site of c-met is achieved by constitutively active Alb-Cre transgene and thus restricted to hepatocytes whereas nonparenchymal liver cells are not targeted. The lack of c-Met expression in hepatocytes was confirmed by PCR analysis and immunohistochemistry  immunohistochemistry and mitotic frequency. A quantitative time course of BrdU incorporation showed that kinetics of DNA synthesis was conserved within the first 36 hr after PH but the peak level was significantly reduced in c-Met mutant livers  represents a key step in chromosome remodeling during cell division  mitosis . These d0-G1 transition of normally quiescent hepatocytes c-Jun, c-Fos and c-Myc between wt/wt;Alb-Cre+/−Met and fl/fl;Alb-Cre+/−Met mice  which allows rapid activation of transcription factors promoting G+/− mice . Also, bafter PH . Nonetheneration .P≤0.01) . In addition, we found downregulation of several genes encoding kinetochore and centromere-associated proteins essential for faithful chromosome segregation during mitotic progression, such as Ndc80, Cenp-e, -g, -h, -k, -i and –q. Among those, Cenpe is a kinesin-like motor protein which accumulates in G2 phase just before the mitosis and required for the efficient recruitment of BUBR1, MAD1 and MAD2 to kinetochores Ect2, ASPM, Nusap1, Kif18a and Stmn1, 3, and 4. Furthermore, the expression of common markers of checkpoint/arrest involved in G1/S and S phase transition, such as Ccne, Ccnd, Gas1, and Dbf4, was also decreased. This was consistent with the observation that the number of S phase cells was reduced in c-Met mutant livers after 36 hr of liver regeneration as cells failed to transit into next S phase providing further support that c-Met deletion has a broad impact on cell cycle progression  . IngenuiP-values . Many ofgression .fl/fl;Alb-Cre+/−Met regenerating livers against a gene set which was previously identified as G2/M phase regulated in synchronized HeLa cells fl/fl;Alb-Cre+/−Met mice at 36–48 hr  to compare the differentially expressed genes in 36–48 hr . Thus, mfl/fl;Alb-Cre+/−Met hepatocytes express a kinase-inactive c-Met receptor which is irresponsive to HGF stimulation The critical role of c-Met in activation of the downstream mediator Erk1/2 during liver regeneration has been established using a liver-specific c-Met conditional knockout mouse model Activation of other members of MAPK family, including a well known regulator of G2/M transition p38 MAPK, appeared to be normal. In addition, we observed only minimal if any changes in phosphorylation of Akt, another important downstream mediator of c-Met pathway. More significantly, there were no differences in the expression of TGFα or phosphorylation of EGFR receptor  suggestifl/fl;Alb-Cre+/−Met mice, we repeated the experiments using regenerating livers from fl/fl;Mx1-Cre+/−Met conditional knockout mice. The results showed that cdc2 and Aurora B expression were similarly repressed at 36–42 hr concordant with the lack of ERK1/2 activation  along with others implicated in spindle organization and biogenesis . In addition, fl/fl;Alb-Cre+/−Met livers displayed downregulation of Egr-1, an immediate early growth response gene shown to be involved in the control of mitotic entry through the spindle assembly checkpoint The role of MAPK/Erk pathway in proliferation of rodent hepatocytes is well known Most significantly, and consistent with lack of Erk1/2 phosphorylation, we found a profound down-regulation of all three classes of serine/threonine kinases that monitor various aspects of mitosis, including Cdk1, Plk1 and two members of Aurora family kinase, Aurora A and B . Among tfl/fl;Alb-Cre+/−Met livers were grown in the continuous presence of EGF. A persistent EGF supplementation in vitro was able only partially rescue the effect of Erk1/2 downregulation in c-Met depleted hepatocytes and restore to some extent the levels of cdc2, Aurora A and B, and Mad2. These results highlight a dominant role for c-Met signaling in maintaining sufficient levels of Erk1/2 activation and suggest that loss of Met/Erk1/2 signaling cascade is responsible for the c-Met knockout phenotype. In conclusion, we report that c-Met function in hepatocytes is required for direct activation of Erk1/2 at the G2/M transition and coordinated control of G2/M transcriptional program.To compensate for the loss of Erk1/2 activation, hepatocyte cultures established from All experimental procedures were approved by the NCI-Bethesda Animal Care and Use Committee (ACUC) under LEC-038. The NCI animal program meets or exceeds requirements of the Public Health Service Policy on Humane Care and Use of Animals and is fully accredited by AAALAC International.fl/flMet;+/−Alb-Cre) and control  mice were described previously. Since deletion of floxed DNA in hepatocytes is time-dependent met deletion was verified by immunohistochemistry. Expression of c-Met was not detectable in fl/fl;Alb-Cre+/−Met hepatocytes and limited to nonparenchymal liver cells  (Sigma-Aldrich) with a 2 day interval. Mx1-Cre-negative littermates (fl/flMet) received equal amounts of poly (I:C) and served as controls. The partial hepatectomy was performed 3 days after the last injection. We used functional test to verify the efficient inactivation of the c-met gene after Mx1-cre-induced recombination. HGF mediated c-Met phosphorylation as well as downstream signaling via Erk1/2 and Akt were completely abolished in hepatocytes isolated from fl/fl;Mx1-c-MetCre+/− mice (To achieve systemic deletion of +/− mice .P<0.05), respectively, reflecting small albeit persistent differences in liver mass , albumin, and triglyceride, were measured using a Reflovet Plus bench top analyzer (Roche) following manufacturer's instructions.Mice received i.p. injections of 150 µg/g 5-bromo-2′-deoxyuridine (BrdU) (Roche Applied Science) 2 hr before sacrifice. Livers were fixed in alcoholic-formalin for 4–6 hr, 5-µm paraffin sections were prepared and stained with ant-BrdU , anti-phosphor-histone H3 Ser10  or PCNA clone PC10  and examined with a Zeiss 510 confocal laser scanning microscope .Whole cell lysates (100 µg) were prepared from frozen tissues using T-PER Tissue Extraction Buffer (Pierce) containing Complete Protease Inhibitor Cocktail (Roche), separated by SDS-PAGE and transferred onto Invitrolon PVDF (Invitrogen). Membranes were blocked in 5% non-fat milk in Tris-buffered saline containing 0.1% Tween 20 for 1 h and probed with specific antibodies listed in Hepatocytes were isolated from 2-month-old mice using a two-step collagenase perfusion 2 transformed. The microarray datasets have been deposited to Gene Expression Omnibus database .Microarrays containing 21,997 long (65-mer) oligonucleotides representing 19,740 unique genes were printed in the Laboratory of Molecular Technology . Total RNA was isolated from frozen liver samples using TRIZOL (Invitrogen) following the manufacture's recommendation. Sample preparation and array hybridization were performed as described t-test P≤0.01) and at least 1.5 fold expression changes between fl/fl;Alb-Cre+/−Met and wt/wt;Alb-Cre+/−Met mice. All selected genes satisfied the criteria for false discovery rate of less than 0.1 which was calculated using “fdrtool” R version 2.11.0 as described The cell cycle progression in the regenerative livers is only a partially synchronized process. Therefore, to identify the biologically relevant gene expression changes, samples at early (30 min-2 hr) and late (36–42–48 hr) time points were pooled together and analyzed as single data sets to minimize the intrinsic variations between different animals while increasing the statistical power of comparison. Differentially expressed genes were selected across the selected time points (early versus late) using bootstrap Data mining was performed using Ingenuity Pathway Analysis tool (Ingenuity Systems Inc.). The significance of each network, function, and pathway was determined by the scoring system provided by Ingenuity. The differentially expressed genes were compared with the genes which were previously identified as G2/M phase regulated in synchronized HeLa cells Figure S1c-met allele (fl/flMet). Genomic DNA was isolated from livers at different age. (B) Indirect immunofluorescence staining of liver sections with c-Met antibody. Nuclear counterstaining was performed with DAPI. Expression of c-Met was not detectable in fl/fl;Alb-Cre+/−Met hepatocytes and limited to nonparenchymal liver cells (arrows). Scale bar, 10 µM. (C) Western blots showing phosphorylation status of c-Met, p42/p44 MAPK and Akt upon HGF stimulation in pimary hepatocytes. Cells were isolated from adult fl/flMet and fl/fl;Mx1-Cre+/−Met livers using a two-step collagen perfusion followed by isodensity centrifugation. After overnight incubation, cells were synchronized by serum deprivation for 2 hrs and treated with 50 ng/ml of rhHGF (PeproTech) for 5–30 min. Note that HGF mediated c-Met phosphorylation as well as downstream signaling via Erk1/2 and Akt were completely abolished in fl/fl;Mx1-Cre+/−c-Met. (D) Representative FACS histograms of DNA content and ploidy distribution in wt/wt;Alb-Cre+/−Met and fl/fl;Alb-Cre+/−Met livers at 2 months of age. Freshly isolated hepatocytes were stained with propidium iodine using the Cell Cycle Test DNA Reagent Kit . Nuclei DNA content was measured using a Becton-Dickinson FACScan flow cytometer and Cell-Quest Sofware. 20,000 events were collected. Results shown are the means ± SE (n = 3 mice per group). (E) Liver/body weight ratios and blood biochemistry. Serum was obtained from wt/wt;Alb-Cre+/−Met and fl/fl;Alb-Cre+/−Met at 2 month of age. Results shown are the means ± SE (n = 3 mice per group). Asterisk indicates statistical significance assessed by Student's t test (P<0.05). M, Marker; AST, Aspartate aminotrasferase, ALT, Alanine aminotrasferase.Selective disruption of c-Met in hepatocytes does not affect postnatal liver growth. (A) PCR analysis of floxed (2.00 MB PDF)Click here for additional data file.Figure S2t test (P is less than at least 0.05).Loss of Met delays recovery of liver mass and function after partial hepatectomy. (A) Serum levels of aspartate aminotrasferase (AST) were indistinguishable in control and Met-deficient mice indicating a similar extent of hepatic injury. (B) Slower recovery of liver weight and reduced levels of albumin (C) and triglyceride (D) in Met-deficient livers. Data are shown as the means ± SE (n = 3−5/group per time point). Asterisks indicate statistical significance assessed by Student's (0.33 MB PDF)Click here for additional data file.Figure S3http://frodo.wi.mit.edu/primer3/). The amplification protocol was as follows: 95°C for 3 min, followed by 40 cycles of 95°C for 15 seconds and 1 minute at 60°C, completed by a dissociation curve to identify false positive amplicons. The relative expression level of each gene was normalized to the corresponding levels at 0 hr and calculated using the formula 2(−ΔΔCt). GAPDH and 18s RNAs were used as endogenous reference. The data are presented as the means ± SD (n = 3). Asterisks indicate statistical significance assessed by Student's t test (P is less than at least 0.05).Temporal profiles of mRNA levels of selected genes during liver regeneration. The results from microarray and the corresponding RT-qPCR analyses are shown in left and right panels, respectively. Oligonucleotide primers were designed using Primer3 v.0.4.0 ((0.40 MB PDF)Click here for additional data file.Figure S4fl/fl;Alb-Cre+/−Met phenotype . (B) The expression values of 48 out of 132 orthologous genes involved in G2/M progression were significantly downregulated in fl/fl;Alb-Cre+/−Met mice at 36–48 hr.Gene set enrichment analysis (GSEA). This analysis was performed to compare gene expression data of regenerating mouse livers with a gene set identified as G2/M phase regulated in synchronized HeLa cells {Whitfield, 2002 #17}. To explore the enrichment of G2/M associated genes, we selected orthologous genes between human and mouse microarrays using HomoloGene database of National Center for Biotechnology Information (NCBI). A total of 132 orthologous genes were present at the G2/M stage. (A) Enrichment of the G2/M gene set in (0.28 MB PDF)Click here for additional data file.Figure S5Western blot analysis of cell cycle-associated genes using nuclear extracts from timed liver samples after partial hepatectomy. Samples were probed by Western blotting using the indicated antibodies.(0.67 MB PDF)Click here for additional data file.Table S1fl/fl;Alb-Cre+/−Met regenerating livers at 36–48 hr .List of genes differentially expressed in (0.20 MB DOC)Click here for additional data file.Table S2List of antibodies.(0.05 MB DOC)Click here for additional data file.


A coordinated response to HIV/AIDS remains one of the 'grand challenges' facing policymakers today. Global health initiatives (GHIs) have the potential both to facilitate and exacerbate coordination at the national and subnational level. Evidence of the effects of GHIs on coordination is beginning to emerge but has hitherto been limited to single-country studies and broad-brush reviews. To date, no study has provided a focused synthesis of the effects of GHIs on national and subnational health systems across multiple countries. To address this deficit, we review primary data from seven country studies on the effects of three GHIs on coordination of HIV/AIDS programmes: the Global Fund to Fight AIDS, Tuberculosis and Malaria, the President's Emergency Plan for AIDS Relief (PEPFAR), and the World Bank's HIV/AIDS programmes including the Multi-country AIDS Programme (MAP).In-depth interviews were conducted at national and subnational levels (179 and 218 respectively) in seven countries in Europe, Asia, Africa and South America, between 2006 and 2008. Studies explored the development and functioning of national and subnational HIV coordination structures, and the extent to which coordination efforts around HIV/AIDS are aligned with and strengthen country health systems.Positive effects of GHIs included the creation of opportunities for multisectoral participation, greater political commitment and increased transparency among most partners. However, the quality of participation was often limited, and some GHIs bypassed coordination mechanisms, especially at the subnational level, weakening their effectiveness.The paper identifies residual national and subnational obstacles to effective coordination and optimal use of funds by focal GHIs, which these GHIs, other donors and country partners need to collectively address. A coordinated response to HIV/AIDS remains one of the 'grand challenges' facing policy makers today . As the Whilst single country studies and broad-brush reviews are starting to reveal the complex relationship between GHIs and efforts to coordinate the HIV/AIDS response ,7, synthAt the global level consensus has emerged about the need to improve coordination of health and HIV-specific programmes -10. SeveDelivering as One added emphasis to the need for better country coordination by outlining a series of reforms to streamline the work of UN agencies operating at country level  ineffective to have multiple coordination structures: the same donor is a member of CCM, member of ICC and is also in the SWAp'. Problems were reported in Ukraine, where multiple national and subnational HIV/AIDS structures exist within a complex, fragmented system of public administrative bodies inherited from the Soviet health system. The study revealed the multiple HIV/AIDS-related structures to have poorly-defined, delineated and overlapping objectives, functions and responsibilities that continue to embrace public sector working practices: their work was neither transparent, nor accountable, with no information about meetings and decisions taken being made public.The studies in Mozambique, China and Ukraine in particular suggest that the multiplicity of parallel national and/or subnational coordination structures have challenged effective governance of HIV/AIDS programmes ,35,37-39In some cases the transience of coordination structures has undermined their effectiveness. In the volatile political environments of Ukraine and Kyrgyzstan, HIV/AIDS coordination structures have been established (and abolished) several times, creating programmatic delays and confusion. Conversely, coordination efforts have benefited from relatively stable, albeit increasingly complex, coordination environments in Mozambique, Zambia and Peru. In Mozambique the CCM secretariat continued to exist as a separate entity, despite integration of the CCM into the SWAp Health Partners Group. In Zambia, the CCM has operated in parallel to the NAC and other national coordination structures ,31,39.Global Fund CCMs were diverse and integrated in different ways and to greater or lesser extents with other country structures, which demonstrates the Fund's evolution since the early years when CCMs were often stand-alone structures and seen as being imposed . The CCMA major goal of HIV/AIDS coordination structures is to promote multisectoral decision making, specifically to involve non-health government departments and nongovernmental actors. Earlier studies ,46 and tNevertheless the studies suggest that despite these developments overall levels of participation and/or engagement of non-health government departments and nongovernmental actors in national and subnational coordination structures remained relatively modest. While no major groups were excluded from membership of national coordination structures in Mozambique and Zambia, in China, Kyrgyzstan, Georgia, Peru and Ukraine non-health government departments were either absent or had marginal engagement; indeed in those countries HIV/AIDS tended to be viewed as a Ministry of Health (MoH) responsibility reflecting the commonly held discourse that HIV/AIDS is a health rather than a broader social issue ,29,32-39In the post-Soviet countries of Georgia, Kyrgyzstan and Ukraine, specialisation within the health system has inhibited interaction between different parts of the system, and between health and non-health departments . UkrainiSimilarly poor coordination between government departments, between different levels of government and poor internal coordination/communication within some government agencies was also reported in China, although the establishment of the CCM was reported as improving government coordination around HIV/AIDS programmes. Additionally, in Kyrgyzstan the position of the national HIV/AIDS coordination structure had hindered attempts at multisectoral decision-making: the structure was relocated from Presidential to MoH level in 2008 ,29. As aWe tried really hard for a long time to make HIV/AIDS problem to be recognised as a social problem in our country. However, if the Secretariat is now by the Ministry of Health, it means that HIV/AIDS became the health problem again.The studies suggest that all three GHIs have created opportunities for CSO involvement in HIV/AIDS programmes through funding their activities, or insisting on their inclusion in CCMs . The Mozambique study reveals that the integration of the CCM within the SWAp increased national level engagement of CSOs and the private sector. Similarly the research in Zambia found that CSOs have begun to play a significant role in district coordination structures, and the World Bank, through the Zambia National Response to HIV/AIDS Project (ZANARA), supported community responses to HIV/AIDS by financing community based organisations, which also participate in District AIDS Task Forces and Community AIDS Task Forces ,31. HoweIn Georgia the CCM membership was described as too large to be manageable. Lead ministries had more than one representative, while other ministries and NGOs were poorly represented: the private sector, religious organisations and education were absent. In order to address this problem the number of CCM members was decreased from 46 to 30 and a rotation principle introduced to manage civil society representation whereby NGOs would elect their representative annually, with two NGOs acting as permanent CCM members. This approach also ameliorated some of the problems of conflicts of interest among NGOs receiving Global Fund grants .However, in common with previous studies and reviews ,22,48, C• Competition for scarce resources at national and subnational level that created distrust between country organisations  and hence a substantial disincentive to meaningful engagement in coordination structures ;• Limited experience among most CSOs of engaging in strategic or political decision making;• Limited financial resources and time to commit to meetings including costs of travelling, and no financial incentives such as per diems and honoraria to encourage attendance (Kyrgyzstan and Ukraine);• Insufficient time to contribute to proposals with tight submission deadlines (Peru);• Government officials at national and subnational level selected CSOs to participate in coordination structures thereby excluding others (China) -35,37,38Unless coordination structures have authority and are seen to be under country ownership, any decisions they make may be ignored potentially leading to poor alignment of GHI and donor programmes with government priorities. The studies explored the extent to which donors were accountable to country coordination bodies and the strength of leadership and political commitment to HIV/AIDS programmes. In Peru and China the studies showed that NACs were able to make decisions and to allocate resources to HIV/AIDS programmes. By comparison national and particularly subnational structures in Zambia, Mozambique, Ukraine and Kyrgyzstan had limited authority to make decisions or allocate resources to HIV/AIDS programmes ,34,35,39An important reason for this was that major donors for HIV/AIDS programmes including PEPFAR continued to set priorities outside national and subnational structures; and their participation in such structures was seen as a formality. Donor interests continued to undermine country ownership and make coordinating multiple aid programmes difficult for countries ,49. The ... most people, when you ask them where they were working, they will tell you that they are working for the [donor] funded project. It's never a Zambian project. So I would like to see a situation where it is... The logo on the vehicle should just say: the Zambian national response to HIV/AIDS and not tell us where the money is coming from.In Zambia and Mozambique the studies found that national coordination structures could not hold the myriad of donors and implementers to account for the effectiveness of their programmes, especially those CSOs that received funding through other channels. PEPFAR and the World Bank participated in NACs in those countries, but PEPFAR recipients in Zambia had limited engagement in subnational coordination structures. Limited decision making and resource allocation powers have been particularly acute within subnational structures, which in practice worked as implementers of local programme determined at the national level rather than as coordination bodies. Donors frequently bypassed such structures. In Zambia government subnational coordination structures, the District AIDS Task Forces, have had a technical/coordination role rather than decision making or resource allocation powers: respondents observed that there was no obligation for GHI-funded NGOs to report to District AIDS Task Forces; they frequently worked to their own priorities and did not participate in them. As a consequence these structures have had very limited control over donor activities and those of international NGOs, and often had minimal information on their activities including how PEPFAR funds were being spent in their districts. Some informants suggested that donor funds were being allocated to programmes which did not coincide with district priorities, leading to service duplication ,31. One One of the challenges when a donor moves into the district, you just see a donor is working there. All they will say is we have been to the Ministry of Health or Education, we got permission and we are working here...The positioning of coordination structures within the wider public administration system has important implications for levels of country ownership and the authority a structure can exercise. An important reason for positioning NACs under the Presidential Office in some African countries has been to give the structures political legitimacy and demonstrate political commitment . In KyrgSimilarly in Ukraine the NAC has had an advisory rather than a decision-making function and met only to agree Global Fund proposals, at which point it was labelled a CCM. Subnational structures had very limited decision making power and minimal influence over local budgets for HIV/AIDS programmes ,35. A re... the status of an advisory institution; that is it doesn't make any decisions... the Coordination Council should help coordination. And this is what they don't do. They meet, review issues, make decisions which are often not implemented.In Zambia, Peru, Ukraine and Kyrgyzstan subnational HIV/AIDS coordination structures were seen as particularly weak and as reinforcing centralised decision-making. In Peru respondents reflected on the limited input from subnational stakeholders in preparing Global Fund grant proposals since the need to draft the proposal rapidly made broad participation and consultation from subnational stakeholders impossible. In Zambia there were mixed views from respondents about whether in practice planning was top-down (from the NAC to the district level) or bottom-up. According to the Zambian National HIV/AIDS Strategic Framework, it was yet to be established how the NAC should communicate with lower level structures and the flow of information to NAC from structures at lower levels was not yet clearly outlined. In Ukraine respondents saw the creation of subnational coordination structures as imposed from the national level and/or international donors, and that their decision-making powers to shape programmes and allocating resources were limited. Regional HIV/AIDS coordination committees were a requirement under the terms of the Global Fund grant, although the grant was not used to fund their establishment or recurrent costs -35. A re... as a whole this system is still bureaucratic, vertical [structures] are created... those coordination councils are created down to the bottom, but everything is like it's used to be. Meetings, conferences, happy reports, everything is done, but the epidemic is spreading...Leadership invested by key members of coordination bodies and commitment of high-level government leaders are important factors in controlling HIV/AIDS epidemics in countries . AlthougThe Georgian CCM benefited from the strong leadership of the First Lady, resulting in improved attendance, coordination between ministries, and expedited decision-making. Kyrgyzstan reported committed leadership in some regional coordination structures, although in practice leadership was vulnerable to rapid turnover of members. Strong leadership was also observed in Mozambique and Zambia. Only in Peru was it reported that weak leadership had undermined the NAC's performance -33,36,39Political commitment is illustrated in different ways. In Peru a 'Declaration of political commitment to HIV/AIDS' raised the profile of the disease, and invoked greater multisectoral commitment than previously. However, no formal policy on coordination or partnership existed, which limited progress. In China the government obligated ministries and local government departments to establish coordination structures and engage with issues of HIV/AIDS. Commitment to coordinated working was found in the Zambian National HIV/AIDS Strategic Framework 2006 - 2010 and the Joint Assistance Strategy; and in Kyrgyzstan a number of government policies explicitly call for multisectoral and CSO engagement in HIV/AIDS control -33,37,38The Ukrainian study revealed variable levels of commitment from local government administrations to HIV/AIDS, which had impaired the effectiveness of coordination structures ,35. HoweLow capacity of secretariats in terms of experience, salaries and equipment, and limited clarity about roles among coordination structure members can undermine the working of these bodies . Putzel In Kyrgyzstan, China and Ukraine, respondents reported that Global Fund funding had engendered better communication and transparency between partners and improved clarity of roles and responsibilities ,35,37,38The Global Fund helped the coordination council understand more clearly and accept international procedures, the procedures of openness, open decision- making, transparency, because the Global Fund influenced indirectly the composition of the National council.A lack of clarity over division of roles and responsibilities among coordination structure members was reported in a number of countries . Poorly defined roles among NAC members in Peru delayed the implementation of the Global Fund grant, and in Zambia roles and responsibilities were ill-defined between the NAC, MoH, other ministries and CSOs, and between various subnational structures and actors ,31. In KThe time of people, who are members of Country Multisectoral Coordination Committee is very 'expensive'. And when I see that the agenda includes discussion of issues related to animal health, and only one of the three issues is related to HIV and my work, I ask myself, do I really need to go to this meeting?Only in Mozambique did the country study suggest that roles were clearly defined among members of national coordination structures, in particular after the SWAp structure was streamlined in 2007.Evidence of limited information flows within and between coordination structures was a key finding in most of the countries, which undermined meaningful exchange between members. While there had been considerable improvements in transparency between subnational actors in Zambia, PEPFAR and NGOs funded by the initiative were unwilling to share information with District AIDS Task Forces, which undermined their authority. However, those CSOs that did participate in these Task Forces were credited with improving communication sharing at district level ,31.formal communication continued to exist at all levels, and there remained a lack of transparency among actors [In Kyrgyzstan limited formal coordination existed at all levels, and in the Ukraine working practices were neither transparent nor accountable. While Kyrgyz stakeholders reported that some local coordination councils fostered improvements in informal information exchange, limited g actors ,29. SpeaAt Country Multisectoral Coordination Committee meetings we cannot possibly get detailed information concerning... what and how much funds have been spent. We asked for this information so many times already, but all our attempts failed. We just receive general reports back...Competition for scarce resources at national and subnational level in Peru, Kyrgyzstan, Zambia, Mozambique and Ukraine was reported as creating distrust between country actors. Nevertheless World Bank HIV/AIDS programmes in Zambia and Mozambique have provided capacity support to the NAC secretariats, and are credited with improving transparency and communications -35,39.The empirical evidence collected in these seven countries provides a kaleidoscope of experience and throws light on country systems and their responses to GHIs. There is huge contextual and historical diversity within and between countries, although what is striking about these findings is that countries with very different contexts shared similar experiences of problematic coordination and the effects of GHIs: findings in Zambia and Mozambique, with generalised HIV/AIDS epidemics and high levels of HIV/AIDS financing, were similar to those in the low and concentrated HIV/AIDS epidemic countries of Europe, Asia and Latin America. In common across the seven countries is the finding that the GHIs - in particular the Global Fund - have had many positive effects on national level coordination. The evidence is that substantial new funding for HIV/AIDS control, for which GHIs can take most of the credit, has created opportunities for multisectoral participation, promoted greater political commitment and increased transparency among most partners.However, refractory problems reported in earlier studies ,46 stillsubnational level that the biggest gap between intent and practice was found in 2006-08. This is a particularly problematic trend. It contradicts the growing emphasis on decentralised health sector decision-making that is seen as strengthening the powers of local-level actors in the formulation and implementation of policies and programmes, thereby increasing responsiveness to local needs [The new knowledge that this cross-country synthesis has begun to generate is that it is at the al needs . The stual needs -35. Indeal needs ,58.Many PEPFAR recipient organisations in Zambia did not participate in subnational coordination structures including District AIDS Task Forces, which consequently had little control over these programmes ,31. SubnGiven these tensions, it is not surprising that coordinated HIV/AIDS programmes remain a distant goal for many countries. These studies suggest that poorly functioning coordination structures undermine programmatic coordination, including weak multisectoral decision making, poor levels of oversight and monitoring and evaluation, poor alignment between GHI and donor programmes and national and subnational level priorities, and implementation problems including delays and confusion, inefficient use of resources and duplication of services. The Global Fund, PEPFAR and the World Bank have made an immense contribution to reducing the burden of HIV/AIDS, especially in sub-Saharan Africa. A clear lesson from these country studies is that GHIs and other donors, as well as national governments, now need to acknowledge and address the residual problems in national level coordination and focus more attention and resources on strengthening subnational coordination, if the gap between intent and practice is to be narrowed.There are a number of limitations of the studies drawn on as part of this analysis. Firstly, much of the data focus on the Global Fund, which is present in all seven countries and has transparent processes, which made data collection easier. Less information on World Bank HIV/AIDS programmes  and PEPFAR (Mozambique and Zambia) reflects difficulties in accessing data, and/or patchy engagement by these GHIs in coordination structures in some countries. It is therefore more difficult to generalise about the effects of the World Bank HIV/AIDS programmes and PEPFAR than about the Global Fund. Secondly, less data are available on subnational coordination than national coordination since subnational interviews were not part of the study design in Peru, Mozambique, and Georgia, although national interviewees commented on subnational coordination in Peru. Thirdly, the findings are based on qualitative interview data. Much less documentary evidence was available to corroborate these data, although country teams endeavoured to triangulate interview data to boost the validity of findings. Finally, while studies explored common themes, there was some heterogeneity across the studies in terms of the precise questions interviewees were asked.The evidence suggests that all seven countries are far from realising the UNAIDS 'Three Ones' principle of one multi-sectoral national body for HIV/AIDS coordination. National as well as subnational coordination structures with a remit for HIV/AIDS are proliferating, and in some countries the multiplicity of parallel coordination structures has challenged the effective governance of HIV/AIDS programmes.GHIs are having some positive effects on HIV/AIDS coordination structures, as well as a number of negative effects: while much has been achieved, particularly at national level, many serious problems remain. For instance GHIs have widened stakeholder participation in coordination structures, although engagement from non-health government departments and civil society remains modest. Country ownership of national and subnational coordination is undermined by the weak decision making authority of many coordination structures and limited or perfunctory engagement among GHIs and other donors, particularly at the subnational level. There is evidence that strong leadership within coordination structures and broad political commitment to coordinated approaches to HIV/AIDS programmes have been improving, although weak secretariat capacity, poorly defined roles and responsibilities among members of coordination structures, limited transparency and communications and competition for scarce resources remain persistent problems undermining effective coordination.Despite the many problems of coordination revealed above, there are several practical lessons stemming from the studies that decision-makers in these and other countries might bear in mind when seeking to strengthen the functioning of national and subnational coordination structures. These include the need to augment the capacity of secretariats of both national and subnational coordination structures through financial and technical support, and to carefully consider how best to position a national coordination structure within the public administration system in order to boost its authority and ability to promote multisectoral working. Financial support for CSOs could promote their effective participation in national and subnational coordination structures by enabling them to attend meetings. Other forms of support for CSOs might also be appropriate such as providing training in strategic or political decision making so that they are better able to engage in coordination meetings and more fully contribute to discussions. Clarity about roles and functions was often missing from the examples presented above reinforcing the need to develop and agree terms of reference for the objectives, functions and working practices of national and subnational coordination structures, and to define the roles and responsibilities of individual members.Several knowledge gaps remain: follow-up research would be especially valuable in helping to better understand how the functioning of coordination structures plays out in the effective coordination of health interventions at the programmatic level, including coordinated service delivery. In particular, further research could help understand the functioning of subnational coordination structures and their effects on programmatic coordination since the evidence at subnational level from these and other studies remains weaker than that at national level. Moreover, at present most evidence is based on qualitative data collection: it will be important to build a stronger evidence base derived from both qualitative, as well as robust quantitative, measures to demonstrate the effectiveness of coordination structures and their effects on programmatic coordination.The authors declare that they have no competing interests.NS led on drafting this article. NS, JA, RB, RB, CC, BC, KC, PM, GM, PN, NR, TS, AW, GW and XZ all participated in the conception, design and execution of the study and analysis and interpretation of data. AH contributed to the analysis and interpretation of data. All authors participated in manuscript writing and have read and approved the final manuscript.


Well over six decades since its first description, the Rheumatoid Factor (RF)—autoantibodies recognizing Fc (constant) portion of IgG through their own Fab (antigen binding variable segments)—is believed to have come of age. Autoimmune pancreatitis is a unique form of pancreatitis, biologically characterized by an elevated serum IgG4 concentration. Given the fact that IgG4 myeloma proteins can act as RF, we initially hypothesized that IgG4 in autoimmune pancreatitis might do likewise, hence potentially contributing to disease pathogenesis. Indeed Western blotting clearly showed that IgG4 binds to IgG1 κ, IgG2 κ, IgG3 κ myeloma proteins, as well as to IgG Fc, in line with a typical RF activity. Further experiments however unraveled the unexpected fact that unlike hitherto known RF, IgG4 does not engage IgG Fc through its Fab, but its very own Fc. These data therefore collectively describe a Novel RF (NRF) in autoimmune pancreatitis. In the future, the relevance of NRF, beyond autoimmune pancreatitis, in both diagnosis/prognosis as well as pathophysiology of autoimmune and other systemic diseases where IgG4's role seems paramount, needs to be systematically assessed. Despite the identification or the generation of numerous natural and/or transgenic/gene deficient animal models of autoimmunity, the translation of the clinical symptomatology and or biological/immunological pathophysiology to man and vice versa has led to few tangible results in the fields of diagnosis/prognosis and/or treatment of human autoimmune diseases infra)Autoimmune pancreatitis is an emerging syndromic entity characterized by several cardinal features distinctive from chronic pancreatitis , in contrast to the original RF Serum samples were obtained from 65 autoimmune pancreatitis patients-54 men and 11 women-aged 38–79 years (median 62.4 years) as well as the following “control” populations: 111 suffering from alcoholic or idiopathic chronic pancreatitis, 96 diagnosed with pancreatic cancer, 40 with autoimmune hepatitis, 39 with primary biliary cirrhosis, 20 with primary sclerosing cholangitis, 13 with systemic lupus erythematosus, 7 with Sjögren's syndrome, 3 with progressive systemic sclerosis and from 130 healthy subjects. All sera were stored at −20°C prior to analysis. All patients with autoimmune pancreatitis fulfilled the revised diagnostic criteria proposed by Japan's Pancreas Society. infra) and developed using enhanced chemiluminescence .Throughout experiments, immunoglobulin samples were resolved on 10 % sodium dodecyl sulfate (SDS)- polyacrylamide gels (PAGE) under reducing conditions followed by transfer onto polyvinylidene difluoride membranes . The blots were incubated with a variety of monoclonal antibodies (1∶1000 dilutions)  and IgG Fc  . The specificity of each immunoglobulin sample was confirmed by Western blotting, where membranes were incubated with the corresponding peroxidase-conjugated antibody for each immunoglobulin isotype. For the totality of experiments the following peroxidase-conjugated second antibodies were used: mouse anti-human IgG1 , mouse anti- human IgG2 , mouse anti-human IgG3 (South Biotech 9210-05), mouse anti-human IgG4 (South Biotech 9200-05) (of IgG1 isotype which reacts to the Fc portion of IgG4), goat anti-human IgA (Zymed 62-7420), goat anti-human IgM (South Biotech 2020-05), goat anti-human IgD  and goat anti-human IgE (Biosource AHI0504). Finally, goat anti-human κ light chain  was used to detect IgG4 F(ab')2.The following human myeloma immunoglobulins were used: IgG1κ, IgG2κ, IgG3κ, IgG4κ, IgA1κ, IgA2κ, IgMκ , IgDκ , IgEλ , IgG F(ab')infra). To confirm the reactivity of the patients' IgG4 to Fc fractions of IgG1, IgG2 and IgG3; a preparation of IgG Fc lacking IgG4 Fc was needed. For such IgG Fc was first absorbed by affinity chromatography where the column was bound with anti-IgG4 antibody . The resultant IgG Fc depleted of IgG4 Fc was then used in Western blotting in addition to intact IgG Fc. IgG4 Fc and IgG4 F(ab')2 were generated through papain In order to use an identical patient serum in all experiments, the autoimmune pancreatitis sera used was indeed a pool derived from all 65 patients (500 µl of each serum×65 = 32.5 ml of pooled serum).The IgG4 concentration of this pooled serum was 755 mg/dl as defined by single radial immunodiffusion . This value was further used to construct a standard curve for ELISA  were coated with human myeloma IgG1κ, IgG2κ or IgG3κ (1 µg/well) proteins. Plates were washed and then blocked with 1% bovine serum albumin (BSA) in phosphate-buffered saline (PBS) containing 10 mmole of ethylene diamine tetra-acetic acid (EDTA). After another wash, serum samples were diluted to 1∶5000 and added. Immobilized complexes were then incubated with peroxidase-conjugated anti-human IgG4 monoclonal antibody in enzyme conjugate stabilizer solution  (1∶2000 dilution). The enzyme bound to the wells was incubated in the dark with tetramethylbenzidine substrate solution . The reaction was halted by the addition of stop solution (TMB One-Component Stop Solution Kirkegaard & Perry Laboratories 50-85-05) and the plate was read at 450 nm. To construct the standard curve, the pooled sera of autoimmune pancreatitis patients were serially diluted from 39 to 10,000 ng/ml of IgG4 and the optical density for each IgG4 value was plotted. The optical density of bound IgG4 concentration in each serum sample was converted to an absolute IgG4 value using this standard curve. The concentrations of standards were expressed as IgG4 values from the pooled patient sera. The linear correlation was obtained for each assay system between the absorbance of IgG4 bound to each IgG subclass and the IgG4 value from the pooled sera. The intra-assay variation was less than 5.0%, and the inter-assay variation less than 7.5%. The HRP-conjugated second antibody reacted minimally to coated myeloma proteins.HLA DRB1*0405-DQB1*0401 haplotype The additive “autoimmune” in “autoimmune pancreatitis” stems from several lines of (indirect) evidence. These include a significant association of susceptibility to the disease with premier immune loci i.e. the de facto RF. In order to do so, we first confirmed the identity of each immunoglobulin myeloma protein used in experiments-IgG1κ, IgG2κ, IgG3κ, IgA1κ, IgA2κ, IgMκ, IgDκ and IgEλ-through reactivity with the corresponding HRP-labeled anti-Ig antibody . These results hence indicate that IgG4 from autoimmune pancreatitis patients is able to bind IgG1κ, IgG2κ and IgG3κ as well as IgG Fc (and perhaps to IgG4κ myeloma protein itself)    in 130 healthy controls as well as patients with other autoimmune diseases  including autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis, systemic lupus erythematosus and Sjögren's syndrome (This Fc-Fc interaction is a novel finding and therefore we have decided to name it Novel RF (NRF) in contrast to the “original” RF hereafter termed Classical RF (CRF). It was now worthwhile to investigate the relevance of NRF syndrome , Table 1syndrome  and thatsyndrome . Given tsyndrome . As depisyndrome , Table 1Although some preliminary reports had suggested the presence of pathogenic autoantibodies against specific pancreatic antigens, de facto RF. However, and surprisingly, IgG4 binding to IgG Fc was due to its Fc, rather than to its Fab. This is formally inconsistent with the simplest definition of an autoantibody that is an immunoglobulin recognizing a self antigen through its variable, antigen binding Fab, and consequently that of a RF i.e. an autoantibody recognizing IgG Fc. Based on this contention, but unwilling to introduce yet a new nomenclature, we propose for this new Ig-Ig interaction to be called Novel RF (NRF) in contrast to the original finding in vitro, did show through domain swapping experiments that the binding of a monoclonal IgG4 RF (isolated this time from a rheumatoid arthritis patient) to IgG is through their respective constant segments and not Fab-Fc Western blot analysis showed that IgG4 did indeed bind to IgG1κ, IgG2κ, IgG3κ and IgG Fc, hence acting, till this stage, as a raison d'être of NRF? Is it beneficial or detrimental to the disease process? Is it limited to IgG4 or could be found for other IgG subclasses or Ig classes? Is it restricted to autoimmune pancreatitis or of more general relevance ? A few clues could be presented here. Firstly, one should be reminded of some structural specificities of IgG4, paramount among which are its inability to engage C1q, hence preventing it to activate the classical complement pathway and dynamic in vivo exchange of Fab arms leading to bispecific antibodies infra). Secondly, there is presently no reason to support for NRF to be limited to autoimmune pancreatitis and given the relevance of IgG4 in a vast array of other disorders, it is logical to assume NRF involvement therein. These include beside systemic manifestations of autoimmune pancreatitis, the following disorders for most prevalent and relevant both in terms of frequency as well as public health implications. They include membranous nephropathy, a prevalent etiology of end-stage renal failure, is characterized by deposit of IgG4 along the epithelia surface of glomerular basement membrane What is the in vivo, several not exclusive situations could be envisioned based on circumstantial evidences. (1) IgG4 Fc-Fc binding may have a pathological role within the inflammatory process, or even induce inflammation per se. An example among may: In a mouse lupus nephritis model, aggregated immunoglobulins react with fibronectin physicochemically; this reaction in turn causes clustering of integrins and internalization of immunoglobulins by glomerular endothelial cells, resulting in fine in glomerular injury in vivo heavy (and light) chain swapping leading for unique biological properties including cross-linking of 2 distinct antigens. This adds to the already mentioned unique properties of IgG4 and may have physiological, pathological as well as therapeutic consequences .In more general terms and despite the fact that is is likely premature to speculate much on the physiological/pathological effects of IgG4 Fc-IgG Fc complexes in vivo with IgG autoantigen in an unprecedented topology. This Fc-Fc interaction departs from the present definition of RF as a Fab-Fc interaction hence the proposed dichotomic nomenclature of CRF vs. NRF. The fact that IgG4 autoantibodies have been universally found in man In conclusion, a sequential set of experiments, aimed initially to confirm the mode of interaction of supposedly RF-like autoantibodies to their cognate autoantigens revealed the unexpected fact that IgG4 autoantibodies interact Figure S1Detailed protocol for generation of IgG fractions.(0.06 MB DOC)Click here for additional data file.


Since its discovery, HIV/AIDS has arguably captured more attention among the Chinese biomedical research community than most other infectious diseases. Traditional parasitic diseases, on the other hand, are perceived as being increasingly neglected. However, it has long been recognized that interactions between HIV and other infective agents, including parasites, influence the health status of people living with HIV/AIDS. This study aimed at systematically reviewing the Chinese scientific literature on HIV/AIDS and parasites between 1986 and 2006 in order to substantiate or refute these claims, and to highlight neglected research areas.Pneumocystis carinii co-infection.Searching the three largest Chinese scientific literature databases, in the China National Knowledge Infrastructure (CNKI) a total of 24,511 citations dealing with HIV/AIDS and 15,398 parasite-specific publications were identified. Wanfang Data and VIP Information (VIP) contained 15,925 and 13,873 entries dealing with HIV/AIDS respectively, while 12,043 and 7,068 hits were scored when searching for parasitological references. The number of publications dealing with HIV/AIDS in China increased exponentially from 6 in 1986 to 3,372 in 2006 whereas the publication activity in the field of parasitology was more erratic and lately started to decline. Epidemiology was the most-reported field of endeavor, accounting for 26.0% and 24.6% of the HIV/AIDS and parasitological literature, respectively, while publications dealing with health education only represented 2.9% and 0.7% of all publications, respectively. The total number of Chinese articles focusing on HIV/AIDS and parasite co-infection was 650, with large year-on-year differences in publication numbers. The single-most frequently studied system was HIV-The present study revealed that in China, the fields of parasitic diseases, especially opportunistic parasitic infections linked with HIV/AIDS, is increasingly neglected. This suggests a need to enhance research in the field of opportunistic parasitic infections and parasitology in general. It usually takes years after an infection with the Human Immunodeficiency Virus (HIV) for Acquired Immunodeficiency Syndrome (AIDS) to develop. The progressing decline and ultimate collapse of immune system functions which are characteristic for AIDS usually result in morbidity and ultimately death due to opportunistic bacterial, viral and parasitic infections and other conditions. Since the first AIDS case was diagnosed in Los Angeles in 1981 ,3.In 1985, the first AIDS case in China was diagnosed at the Beijing Union Medical College Hospital . OriginaThe Chinese research community soon became involved in HIV/AIDS research and quickly expanded its activities in the field as official recognition and encouragement were offered, and more dedicated funds were made available. In line with these developments and wider trends in research, the number of scientific publications about HIV/AIDS in China multiplied both domestically and in the international scientific literature. Today, the available domestic Chinese literature is diverse and broad in scope and approaches, and the growth in the number of publications is unabated.Parasitic infections are still common in many regions and populations across China, and represent a lasting public health challenge. The results of the "National sampling survey of the current situation of major human parasitic diseases in China", which had been implemented between 2001 and 2004 throughout the country, suggest that the overall prevalence of common soil-transmitted helminth infections across China was 21.7%, and that 7.9% of the targeted population segments tested sero-positive for toxoplasmosis. Other parasites were also commonly encountered  and addi3 [Mycobacterium tuberculosis and viral hepatitis.It has long been recognized that interactions between the HI virus and the host facilitate the colonization of the latter by other infective agents, including parasites. This negatively impacts on the health status and outlook of people living with HIV/AIDS. With the progressing development of AIDS, especially once CD4+ T-lymphocyte counts have fallen below 200/mm3 -11. A si3 ,13. HoweThe objectives of the present study were i) to systematically review the peer-reviewed scientific literature on HIV/AIDS, parasites and co-infections published in China between 1986 and 2006; ii) to analyze publication dynamics and classify the identified communications according to research topics; and iii) to highlight neglected research areas.More than 10 online scientific literature databases are currently available in China, covering the entire spectrum of the Chinese biomedical literature. The three most comprehensive and widely used databases for searching and retrieval of Chinese scientific publications are the China National Knowledge Infrastructure (CNKI), VIP Information (VIP) and Wanfang Database ,15. AmonPneumocystis', 'cryptozoite', 'Toxoplasma', 'Cryptosporidium', 'amoeba', 'malaria' and 'Leishmania'.References in CNKI, VIP and Wanfang were identified using the following approach. First, scientific publications dealing with HIV/AIDS were identified using the search term The retrieved citations were imported into the literature management software EndNote version 9.0 . After manually removing duplicates, comments on previous publications, solicited articles, conference abstracts and notifications, temporal publication patterns were established by stratifying the identified articles by publication year, and detailed analyses of the focus and content of the identified publications were performed. The categories 'case report', 'diagnosis', 'epidemiology', 'health education', 'immunology & molecular research', 'intervention & control', 'reviews', 'clinical therapy & drug resistance' and 'vaccine development' were used to characterize the research field from which individual publications hailed.Using identical search terms and limitations for querying the three largest Chinese databases for scientific literature, the highest number of hits resulted from querying CNKI. In this database, a total of 24,511 references dealing with HIV/AIDS and 15,398 publications about parasites were identified in the specified time period. In Wanfang database, 15,925 (HIV/AIDS) and 12,043 (parasites) entries were highlighted, respectively. The least number of hits resulted from querying the VIP database where 13,873 HIV/AIDS-related and 7,068 parasite-specific citations could be identified.Screening the retrieved citations from the CNKI for duplicates, comments etc. resulted in the removal of 486 (HIV/AIDS) and 487 (parasites) references. Hence, a total of 24,025 citations dealing with HIV/AIDS and 14,911 parasite-specific publications were available for further analysis. As shown in Figure The stratification of the identified publications dealing with HIV/AIDS and parasites, respectively, by their research emphasis is displayed in Table A total of 388 publications dealing with HIV and parasite co-infections were identified among the Chinese scientific literature published between 1986 and 2006, and met the inclusion criteria for further analysis. Figure Pneumocystis carinii co-infection was the single-most researched system, accounting for 30.9% of all publications. The second-most frequently studied associations were those involving Cryptosporidium and Toxoplasma, accounting for 12.6% and 11.8% of the total number of publications, respectively. To put the number of communications reporting novel research findings into perspective with other types of information generation, it must be noted that almost half of the total number of publications were reviews (48.2%). The most commonly reported original data were case reports which represented 14.4% of all publications, followed by issues related to diagnosis (12.9%) and therapy (10.6%). The smallest category was epidemiological investigations, representing only 6.4% of the identified titles.A stratification of these 388 publications on HIV/AIDS and parasite co-infections is displayed in Table P. carinii, Cryptosporidium and Toxoplasma were analyzed in further detail. The prominence of different research fields varied between the three systems. Among 120 Chinese titles reporting HIV and P. carinii co-infections between 1986 and 2006, reviews, diagnosis, case reports, therapy and epidemiological investigations represented 29.2%, 25.8%, 21.7%, 15.8% and 3.3% of the total, respectively. The 49 publications dealing with the HIV-Cryptosporidium system fell into the categories reviews, diagnosis, therapy, epidemiological surveys and case reports in decreasing order of prominence. Similarly, the 46 titles on HIV and Toxoplasma co-infections comprised reviews as the most prominent category, followed by case reports, epidemiological surveys, therapy and diagnosis.Chinese communications on HIV and parasite co-infections focusing on P. carinii among HIV positives were most prominent, accounting for 62% of all titles in this category. The diagnosis of Cryptosporidium and Toxoplasma co-infections represented 18% and 4%, respectively.Overall, the diagnosis of HIV and parasite co-infections represented the largest category of original data and accounted for 12.9% of all publications on HIV-parasite co-infections between 1986 and 2006. Publications dealing with the diagnosis of Following the official acknowledgement of local HIV transmission in China by relevant authorities, it soon became evident that the disease was of potentially huge public health significance for the world's most populous country. The Chinese research community started to pay attention to the emerging threat even when the initial neglect and downplay from public institutions still continued, and little means for research were available. Soon after the acknowledgement of the looming crisis by high-ranking officials, resources for research became more readily available, and the topic also caught the attention of the general public. This change in attitudes is well reflected in the evolution of publication numbers. The absolute numbers of publications on HIV/AIDS in China as well as the steep increase over time provide firm evidence that this research field became one of the focus topics of Chinese public health and infectious disease research. It is also likely that the observed trend will continue in the foreseeable future since the challenges posed by HIV/AIDS to public health in China are unlikely to wane any time soon. Thus, it appears highly significant that the government promoted this research field, only after which it started to flourish. China has a tradition of tightly overseeing and steering research and promoting research fields which have been officially identified as top priorities. Thus, the government should continue to foster multi-sectoral and international collaboration and encourage innovation in research , withoutOver the 20-year's history of HIV/AIDS research in China, a diverse approach covering a wide range of basic and applied research questions, from vaccine development through clinical medicine to health education, has developed. The unfolding of the domestic research field partially followed international trends, but there are also differences. In the domestic Chinese literature, the relative contributions of the different research avenues to the overall corpus of HIV/AIDS-specific literature are generally comparable to that of the dedicated international literature . The onlClonorchis sinensis infection in 1990 with that at the beginning of the 21st century. It found that the prevalence had significantly increased in Guangdong and Jilin provinces as well as Guangxi Zhuang Autonomous Region, with reported increases of 182%, 630% and 164%, respectively [Angiostrongylus cantonensis [Compared to the dynamic development of HIV/AIDS research, the publication activity on parasitologic topics in China appears more restrained. Over the 20-year period investigated here, a modest increase followed by a stable period and, later, a decreasing trend was found. One explanation might be the ongoing epidemiological transition, i.e. a broader shift from infectious, including parasitic, diseases to chronic non-communicable diseases as the main public health problem in China. This phenomenon can be observed globally in the wake of broad socio-economic development and is arguably more pronounced in China with its rapid development and lifestyle changes over the last 30 years. Viewed from this perspective, it is reasonable that parasitology and parasitic diseases received less attention in relative and even absolute terms after having been controlled in many areas across China. However, parasitic diseases are far from controlled across much of China . Even cotonensis . Thus, ctonensis . This neIn HIV/AIDS research, high numbers of studies were and continue to be carried out in China by local researchers and research consortia, and many interesting results were obtained. However, most research findings were published in domestic journals only available in Chinese, thus greatly hampering the exchange with the international scientific community. For most non-Chinese researchers the Chinese literature is not accessible due to language and technology barriers , incurring scientific and economic costs which await quantification but might be substantial. Some issues of concern are study duplications and hence uneconomic resource allocation, and missed potential for significant insights which can result from integrating findings originating from different studies conducted at various locations and under different conditions. Especially in rural areas, where local healthcare professionals generate a huge amount of relevant data, a notable shortage of funds, infrastructure and training as well as communication skills hamper improvements in the quality of scientific research. The most that can usually be generated under such circumstances are studies on clinical diagnosis, treatment and individual case reports. It has previously been concluded that the domestic Chinese literature were a huge treasure trove which should be explored and opened up to the international community ,24,25. IThe number of publications dealing with HIV/AIDS and parasite co-infections published in China is limited. The majority of these articles are reviews (often of studies conducted outside China) and case reports. Even basic research questions still await a definitive answer in China. For example, there are no comprehensive data on the overall prevalence of different parasitic infections among AIDS patient in China. This is especially troubling if seen against the backdrop of the high fraction of HIV infections stemming from poor rural regions of China, and the wide variety of parasites which still thrive in these areas. A distinct subgroup among the global HIV cohort which is prominent in China is commercial blood donors. In China, HIV infections among blood donors occurred at least up to 1994 due to risky procedures, one of the worst public health scandals of modern China . The majHIV/AIDS research in China has ballooned over the last 20 years following focused government attention and steep increases in funding. By contrast, parasitology lost ground in relative but also absolute terms, and publication numbers are declining. The structure of the published research suggests a predominance of descriptive and repetitive studies, with relatively little innovation. The large number of publications only available in Chinese suggests missed chances for scientific advancement due to ignorance of research results and duplication of studies, indicating an uneconomic use of overall research funding. Research on parasitology in general and on HIV/AIDS and parasite co-infections in particular should be strengthened in order to better reflect the public health relevance of the respective infections. The exchange between the Chinese and the international research community should be promoted by encouraging the publication of significant findings by Chinese scientists in international journals, the publication of major review articles in the respective other language, and by the promotion of English abstracts and articles in Chinese scientific journals and their improved accessibility for foreign researchers.AIDS: Acquired Immunodeficiency Syndrome; CNKI: China National Knowledge Infrastructure (a literature database); HIV: Human Immunodeficiency Virus; VCT: Voluntary Counseling and Testing; VIP: VIP Information (a literature database).The authors declare that they have no competing interests.LGT conceived the study, carried out data collection and analysis and drafted the manuscript. PS conceived the project, assisted in the interpretation of the results and revised the manuscript. JXC and SHC conceived the project and provided technical support for data collection and analysis. XNZ conceived the study and revised the manuscript. All authors read and approved the final manuscript.


Africa continues to bear a disproportionate share of the global HIV/AIDS, tuberculosis (TB) and malaria burden. The development and distribution of safe, effective and affordable vaccines is critical to reduce these epidemics. However, conducting HIV/AIDS, TB, and/or malaria vaccine trials simultaneously in developing countries, or in populations affected by all three diseases, is likely to result in numerous ethical challenges.Convergent Ethical Issues in HIV/AIDS, TB and Malaria Vaccine Trials in Africa in Durban, South Africa on the 10-11 February 2009.In order to explore convergent ethical issues in HIV/AIDS, TB and malaria vaccine trials in Africa, the Ethics, Law and Human Rights Collaborating Centre of the WHO/UNAIDS African AIDS Vaccine Programme hosted a consultation on the Key cross cutting ethical issues were prioritized during the consultation as community engagement; ancillary care obligations; care and treatment; informed consent; and resource sharing.The consultation revealed that while there have been few attempts to find convergence on ethical issues between HIV/AIDS, TB and malaria vaccine trial fields to date, there is much common ground and scope for convergence work between stakeholders in the three fields. Africa continues to bear a disproportionate share of the global burden of HIV, TB and malaria. There are 22 million people living with HIV in sub-Saharan Africa, 67% of the 33 million people living with HIV globally . An estiDespite the variety of prevention, treatment and care initiatives in existence, significant challenges in curbing these diseases remain. The development and distribution of vaccines that are safe, effective and affordable is critical to reduce disease and mortality from HIV/AIDS, TB, and malaria. The geographic overlap, shared risk factors and possibility for co-infection between HIV/AIDS, TB, and malaria mean that these diseases can no longer be considered in isolation. International cooperation and the development and testing of multiple vaccines simultaneously, are required to curb these epidemics.The last few years have seen significant progress in the development of vaccines against HIV/AIDS, TB, and malaria. Despite several setbacks, the search for a safe and effective HIV/AIDS vaccine continues. There are presently HIV/AIDS vaccine trials being conducted in four African countries namely Uganda, Kenya, Tanzania and South Africa . Proof-oConducting vaccine trials in developing countries is ethically complex and requires careful consideration of several key issues . HIV/AIDConvergent Ethical Issues in HIV/AIDS, TB and Malaria Vaccine Trials in Africa in Durban, South Africa on the 10-11 February 2009. This meeting aimed to bring together key stakeholders from HIV/AIDS, TB, and malaria vaccine initiatives to explore the convergent ethical issues in the three fields and to promote and impact cooperation, networking and resource sharing in Africa. Participants included sponsors, investigators, research ethics committee members and community representatives working in the fields of HIV/AIDS, TB and malaria vaccine research. This paper outlines the key convergent ethical issues and recommendations identified during the consultation.In order to explore convergent ethical issues in HIV/AIDS, TB and malaria vaccine trials in Africa, the Ethics, Law and Human Rights Collaborating Centre of the WHO/UNAIDS African AIDS Vaccine Programme hosted a consultation on the Using an informal ranking exercise, the key cross cutting ethical issues in HIV/AIDS, TB, and malaria vaccine research in Africa, were roughly prioritized by stakeholders in the consultation as follows,: 1) Community engagement; 2) Ancillary care obligations; 3) Care and treatment; 4) Informed consent; 5) Resource sharing. These are detailed below, with recommendations for each.Community engagement approaches appear to be better developed in HIV/AIDS research than in research relating to TB and malaria. HIV/AIDS research typically includes broad community education campaigns  and community consultation regarding specific studies. Conversely, TB related trials have, in general, not included such community involvement and preparation for research. This might be argued to be a result of the global advocacy around HIV/AIDS . HIV/AIDS advocacy in North America changed human rights globally. HIV/AIDS became more than a health-related issue and is now also a political issue. Despite this difference, stakeholders agreed that relationships with communities are an important ethical concern for all three fields and that community engagement needs to move beyond the tokenistic involvement of the community and towards more power sharing relationships. Community engagement is a critical component in clarifying reasonable expectations regarding study results  and care, for managing issues around the therapeutic misconception and for ensuring that procedures are culturally sensitive. Community engagement is also increasingly desired and required when considering at what stage in trial planning to involve communities and how best to do so.It is recommended that a science of community engagement be encouraged , where sCareful advanced planning on which ancillary diseases will be treated is essential -20. Ancia priori partnerships be forged and formalized between local health service providers and researchers to develop a referral system for ancillary care prior to trial initiation so that participants can be referred for ancillary care, when appropriate.It is recommended that the provision of ancillary care be carefully considered during protocol development, in consultation with host communities, and that responsibility for investigative and ancillary care procedures is effective, clearly differentiated and performed by designated or different members of the trial staff. It is also recommended that Defining the standard of care and treatment to be provided when a participant becomes infected with the pathogen under study is a contentious issue. While TB and malaria are curable and treatments are relatively inexpensive and readily available, HIV/AIDS remains incurable and life-long treatment is required. Standards of care and treatment, particularly for HIV/AIDS, are usually much higher in sponsor countries (Europe and North America) and sponsors may require this higher standard in developing country settings when it is not the norm for the local population. This issue is a particular concern for multi-centre studies where the standard of care and treatment may vary across centres, or across participants and non-participants within a host country. Regardless of which standard is implemented, research sites generally provide better care than that which is provided by the health systems in the host country. Sustainability of this 'higher' standard of care and treatment beyond the duration of the study is a challenge faced by many investigators.Clear boundaries regarding care and treatment obligations need to be delineated. In contexts where referral systems are well-developed and investigators can refer participants, follow-up is crucial as the ethical responsibility to participants does not end once referrals are made. Complex decisions on how care and treatment obligations should be met should be trial specific and made through an open and participatory process before a trial begins ,23. The Factors such as poor health delivery systems, poverty, the therapeutic misconception and illiteracy may hinder the process of obtaining truly informed consent. In malaria and TB vaccine research, where target populations are often children, extra care and effort are needed on the part of research teams when obtaining proxy consent from parents/guardians, especially in cases where the parents may themselves be minors. Sponsors often require long, detailed and overly legalistic consent forms to be administered. These forms are intimidating and challenging to administer due to low levels of literacy among the majority of volunteers. As such, understanding may be limited. Therefore, local investigators argue for short, simplified versions of consent forms. Changing consent form templates involves significant power dynamics between sites and sponsors, and some sponsors may be unwilling to relinquish this. The debate about length, detail and usefulness of consent forms may be seen as a function of over-regulation of the clinical trials industry. Studies are often not approved if they do not include lengthy consent forms, regardless of their appropriateness. This may be reflective of assumptions that mere compliance with regulations regarding what to include in consent forms is equal to meeting certain ethical standards, which is not necessarily the case.Sharing of simple and effective consent form templates and procedures across disease areas is recommended  as is thCollaborative relationships between researchers and research centres, working within, and across, each of the disease entities is critical in promoting infrastructural and capacity development in developing countries, and in responding to ethical challenges. Working in pathogen-specific silos is an artifact, as TB, HIV/AIDS and malaria are related and are often co-morbid, and communities may want attention paid to the spectrum of diseases affecting them. Attempts to diversify across fields should acknowledge that working in silos does tend to serve donor priorities and thus researchers' personal career interests, particularly in light of competition for funding. Sharing of resources and capacity across disease areas may however help to avoid wasting capacity,  which has been established in research teams, between trials, and when candidate products are unavailable. Potential barriers to resource sharing include institutional separation, ignorance of each field's history and unique challenges, constrained resources and different timelines. Potential enablers include identifying common platforms of issues and developing models and tools that can serve more than one initiative. The principal advantages are the maximisation of finite resources and energies, and the promotion and retention of local capacity.Given the limited resources and capacity in Africa, it is recommended that where capacity has been developed, and where sophisticated infrastructure has been put in place, within a disease entity, this should be shared within and across disease entities in order to further the objective of addressing health and development at a broader level. This would include the sharing of training resources  across disease entities. This would also enable the retention of scarce skills and reduce 'brain drain' from the developing world.The consultation revealed that while there have been few attempts to find convergence on ethical issues between HIV/AIDS, TB and malaria vaccine trial fields to date, there is much common ground and scope for convergence work between stakeholders in the three fields. Based on recommendations made during the consultation, a steering committee has been established and possible funding sources have been identified to move the convergence agenda forward. Additional activities recommended to move the convergence agenda forward include: An empirical study of how key stakeholders in HIV/AIDS, TB and malaria vaccine trials would prioritise convergent ethical issues; the documentation of the existence of disease specific 'silos'; a study of knowledge differences and barriers to convergence between the three fields; and the hosting of a larger forum, with better representation from the key stakeholders and decision makers in the three fields, to debate these issues further.AIDS: Acquired Immune Deficiency Syndrome; HIV: Human Immunodeficiency Virus; TB: Tuberculosis; UNAIDS: Joint United Nations Programme on HIV/AIDS.The authors declare that they have no competing interests.DW chaired the consultation and NM, JK and ZE rapporteured the consultation to which the manuscript refers. All authors drafted the manuscript and read and approved the final manuscript.NM is the Project Manager of the African AIDS Vaccine Programme's Ethics, Law and Human Rights Center, School of Psychology, University of KwaZulu-Natal.DW is a Professor at the School of Psychology, University of KwaZulu-Natal, South Africa and the Principal investigator of the South African Research Ethics Training Initiative and the WHO-UNAIDS African AIDS Vaccine Programmes' Ethics, Law and Human Rights Centre, School of Psychology, University of KwaZulu-Natal.JK and ZE are researchers at the South African AIDS Vaccine Initiative's HIV AIDS Vaccine Ethics Group, School of Psychology, University of KwaZulu-Natal.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6939/11/3/prepub