The need for a telemedicine strategy for Botswana? A scoping review and situational assessment.

BACKGROUND: Health, healthcare, and healthcare system problems within the developing world are well recognised. eHealth, the use of Information and Communications Technologies (ICT) for health, is frequently suggested as one means by which to ameliorate such problems. However, to identify and implement the most appropriate ehealth solutions requires development of a thoughtful and broadly evidence-informed strategy. Most published strategies focus on health informatics solutions, neglecting the potential for other aspects of ehealth (telehealth, telemedicine, elearning, and ecommerce). This study examined the setting in Botswana to determine the need for a telemedicine-specific strategy. METHODS: A situational assessment of ehealth activities in Botswana was performed through a scoping review of the scientific and grey literature using specified search terms to July 2018; an interview with an official from the major mhealth stakeholder; and benchtop review of policies and other relevant Government documents including the country's current draft eHealth Strategy. RESULTS: Thirty-nine papers were reviewed. Various ehealth technologies have been applied within Botswana. These include Skype for educational activities, instant messaging (WhatsApp for telepathology; SMS for transmission of laboratory test results, patient appointment reminders, and invoicing and bill payment), and robotics for dermatopathology. In addition health informatics technologies have been used for surveillance, monitoring, and access to information by healthcare workers. The number of distinct health information systems has been reduced from 37 to 12, and 9 discrete EMRs remain active within the public health institutions. Many infrastructural issues were identified. A critical assessment of the current draft ehealth strategy document for Botswana showed limitations. Many telemedicine services have been introduced over the years (addressing cervical cancer screening, teledermatology, teleradiology, oral medicine and eye screening), but only one project was confirmed to be active and being scaled up with the intervention of the Government. CONCLUSIONS: Botswana's draft 'ehealth' strategy will not, in and of itself, nurture innovative growth in the application of telemedicine initiatives, which currently are fragmented and stalled. This lack of focus is preventing telemedicine's recognised potential from being leveraged. A specific Telemedicine Strategy, aligned with and supportive of the pre-existing ehealth strategy, would provide the necessary focus, stimulus, and guidance.

Health needs and eHealth readiness assessment of health care organizations in Kabul and Bamyan, Afghanistan.

This study assessed the need and readiness of health care institutions in Kabul and Bamyan, Afghanistan for successful implementation of information and communication technology in health care (eHealth). A mixed methods design was adopted at 2 institutions in the Aga Khan Development Network in Afghanistan: the French Medical Institute for Children in Kabul and Bamyan Provincial Hospital, Bamyan. Information for the needs assessment was obtained from interviews and focus groups and eHealth readiness was assessed using a validated survey tool. The needs of institutions in the Aga Khan Development Network in Afghanistan were categorized as follows: provision of care needs; learning needs; and information management needs. eHealth readiness on average was lower in Bamyan compared with Kabul in all areas of the readiness assessment. Other institutions in Afghanistan may benefit from adopting the model of needs and readiness assessment used for Aga Khan Development Network institutions.

The structure of erythrocyte membranes studied by freeze-etching. II. Localization of receptors for phytohemagglutinin and influenza virus to the intramembranous particles.

The distribution of specific glycoprotein receptors on the external surfaces of red cells was mapped, by the freeze-etching technique, to determine if the receptors coincided with the underlying 75-A intramembranous particles. Phytohemagglutinin, ferritin-conjugated phytohemagglutinin, and influenza virus were used as labeling agents since they can be seen by freeze-etching techniques and each reacts with a different site on the same glycoprotein molecule. The distribution of these labels was studied on intact human red cells, isolated ghost membranes, and trypsin-treated ghost membranes. The results show that the receptors for these labels are distributed uniformly over the surfaces of normal red cell membranes in the same apparent distribution as that of the 75-A particles within the membrane. The association between the external receptors and the underlying particles is especially evident when trypsin-treated ghost membranes are labeled: the labeled receptor sites and the intramembranous particles both form sharply defined, reticulated networks, which overlap. These results provide further support for the idea that membrane-bound glycoproteins are oriented so that their carbohydrate-rich segments, which bear the antigenic sites and receptors, are exposed to the external medium, while hydrophobic segments of the same molecules interact with lipids, and possibly other proteins, to form the intramembranous particles.

Regulation of the terminal event in cellular differentiation: biological mechanisms of the loss of proliferative potential.

Human plasma has been demonstrated to contain factors that induce the sequential expression of nonterminal and terminal adipocyte differentiation in 3T3 T mesenchymal stem cells. We now report the development of methods for the isolation of purified populations of nonterminally differentiated cells and terminally differentiated cells, and we show that it is possible to experimentally induce transition from the nonterminal to the terminal state of differentiation. With this model system it is therefore now possible to examine the biological and molecular processes associated with the terminal event in differentiation, i.e., the irreversible loss of proliferative potential. In this regard, we demonstrate that transition from the nonterminal to terminal state of differentiation is a complex metabolic process that consists of at least two steps and that this process can be triggered by pulse exposure to an inducer for approximately 12 h but that approximately 24-48 h is required for the process to be completed. The data also establish that induction of the terminal event in differentiation requires protein synthesis but not RNA and DNA synthesis. These and additional results suggest that loss of proliferative potential associated with the terminal event in cellular differentiation is a distinct regulatory process, and we suggest that defects in this regulatory process may be of etiological significance in the pathogenesis of specific human diseases, especially cancer.

Differentiation modulates the balance of positive and negative Jun/AP-1 DNA binding activities to regulate cellular proliferative potential: different effects in nontransformed and transformed cells.

Differentiation of 3T3T cells into adipocytes results in the progressive repression of growth factor responsiveness. This is associated with the transcriptional repression of the inducibility of c-jun and junB expression by serum. In contrast, differentiation of SV-40 large T antigen-transformed 3T3T cells (CSV3-1) does not repress growth factor responsiveness nor c-jun or junB inducibility even though CSV3-1 cells can differentiate into adipocytes. To better explain these observations, we have studied compositional changes in AP-1 DNA binding activity attributed to c-Jun, JunB, and JunD during the differentiation process in 3T3T and CSV3-1 cells. The results show that in nontransformed 3T3T cells, differentiation represses AP-1 DNA binding activity via a proportionate downregulation of c-Jun, JunB, and JunD. In contrast, in CSV3-1 cells, AP-1 DNA binding activity increases twofold during differentiation, which is accounted for by an increase in JunD with no change in c-Jun and JunB. If c-Jun and JunB serve as positive regulators and JunD serves as a negative regulator for cell proliferation as suggested by previous studies, the repression of JunD expression in differentiating CSV3-1 cells should be mitogenic because decreasing JunD/AP-1 DNA binding activity would allow c-Jun/AP-1 and JunB/AP-1 DNA binding activities to be dominant. The results confirm this prediction showing that antisense junD oligodeoxyribonucleotides are mitogenic for differentiating CSV3-1 cells whereas antisense c-jun and junB inhibit mitogenesis. These data support the conclusion that differentiation can regulate cellular proliferative potential by modulating the balance of positive and negative Jun/AP-1 DNA binding activities in distinct ways in nontransformed and transformed cells.

Coupling of proadipocyte growth arrest and differentiation. I. Induction by heparinized medium containing human plasma.

The differentiation of proadipocytes in vitro typically required prolonged culture of cells as a high density in high concentrations of serum and added hormones. With such culture conditions it is difficult to design experiments to determine the mechanisms that control the differentiation process. We now describe the rapid and parasynchronous growth arrest and differentiation of low density murine proadipocytes in heparinized medium containing only human plasma. When low density cells are cultured under these conditions, growth arrest at a distinct state in the G1 phase of the cell cycle occurs within 2 d and the differentiation of 80-100% of the cell population occurs within 4 d thereafter. The factors in human plasma which promote growth arrest and differentiation are heat labile and can be separated by barium adsorption. In the following paper we have used these methods to show that there are five separate phases which regulate the coupling of proadipocyte growth arrest and differentiation. The data reported in this paper establish that: (a) high cell density and extensive cell-to-cell contact are not required for adipocyte differentiation, (b) prolonged culture is not required for adipocyte differentiation, and (c) high concentrations of serum and/or added hormones are not required for adipocyte differentiation.

Loss of proliferative potential during terminal differentiation coincides with the decreased abundance of a subset of heterogeneous ribonuclear proteins.

The decrease in abundance of a subset of highly conserved basic nuclear proteins is established to correlate with the loss of proliferative potential in association with the process of terminal differentiation in murine mesenchymal stem cells and human keratinocytes. These proteins, designated P2Ps for proliferation potential proteins, have apparent molecular masses of 30-40 kD, are associated with the 30-40S substructures of nuclear hnRNP complexes, and are recognized by antibodies made against core proteins of hnRNP particles. They also share an epitope in common with heat shock protein-90 (hsp90) and are recognized by two mAbs against hsp90. Two-dimensional electrophoretic Western blots furthermore show that P2Ps make up a subset of hnRNP proteins. Cells that possess these proteins express the potential to proliferate whether or not they are traversing the cell cycle. These include rapidly growing cells, reversibly growth-arrested cells, and nonterminally differentiated cells. In contrast, cells that have irreversibly lost their proliferative potential, such as terminally differentiated cells, show a marked reduction in the abundance of P2Ps as determined by immunodetection on Western blots. A correlation, therefore, exists between the presence of this subset of nuclear proteins and the proliferative potential in two cell types. These results raise the possibility that as a subset of hnRNP proteins, P2Ps may mediate posttranscriptional control of the processing of specific RNAs required for cell proliferation.

Coupling of proadipocyte growth arrest and differentiation. II. A cell cycle model for the physiological control of cell proliferation.

Experimental evidence is presented that supports a cell cycle model showing that there are five distinct biological processes involved in proadipocyte differentiation. These include: (a) growth arrest at a distinct state in the G1 phase of the cell cycle; (b) nonterminal differentiation; (c) terminal differentiation; (d) loss of the differentiated phenotype; and (e) reinitiation of cell proliferation. Each of these events is shown to be regulated by specific human plasma components or other physiological factors. At two states designated GD and GD', coupling of growth arrest and differentiation is shown to occur. We propose that these mechanisms for the coupling of growth arrest and differentiation are physiologically significant and mimic the regulatory processes that control stem cell proliferation in vivo.

Plasma membrane vesiculation: a new technique for isolation of plasma membranes.

Monolayer cell cultures of macrophages, monocytes, myoblasts, and density-inhibited and transformed fibroblasts form and release cell surface membrane vesicles following exposure to formaldehyde, related low-molecular-weight aldehydes, and disulfide blocking agents. Vesicles have a unique composition of proteins and lipids. They show enrichment of cholesterol and sphingomyelin content and a seven-to tenfold enrichment of 5'-nucleotidase activity. Vesicles also contain intramembranous particles and show a trilamellar unit membrane and no ultrastructural evidence of contamination with other cytoplasmic organelles. The technique is proposed as a novel method for isolating plasma membrane vesicles from cells in culture.

Faecal sludge in Accra, Ghana: problems of urban provision.

Urban on-site sanitation services present challenges for emptying, transporting, disposing and treating faecal waste. Transfer stations can be used by household-level emptiers to safely dispose of faecal sludge, but they rarely exist. Accra's use of transfer stations has provided an opportunity to research their functioning, as part of broader faecal sludge management arrangements. The paper discusses the benefits offered by use of transfer stations, as well as reasons currently limiting their operation. While costs associated with operating and emptying these stations are passed to householders, an illegal sector thrives offering lower cost emptying services, typically with disposal of faecal sludge directly into the environment. At present, bucket latrines offer sanitation services to low-income households unable to afford higher service levels, such as septic tanks. The local government aims to phase-out all bucket latrines by 2010, but affordable alternatives have not been found. Where limited access to land inhibits investment in permanent facilities, families may abandon household sanitation altogether. The paper concludes that correct use of transfer stations can provide improvements for existing faecal sludge management and reduce indiscriminate dumping. They must be made available to all workers, through effective public-private arrangements for ownership and operation.

Cyclic adenosine 3',5'-monophosphate-independent phosphorylation of externally oriented plasma membrane proteins in transformed and nontransformed mouse embryo cell lines.

Thirty transformed and nontransformed mouse embryo cell lines were analyzed to determine if increased plasma membrane phosphorylation is a characteristic of all transformed cell lines. Nontransformed, virally and chemically transformed, and smooth surface-transformed cell lines derived from Swiss, BALB/c, AKR, or C3H mice were studied. Cyclic AMP-independent phosphorylation of plasma membrane proteins was comparable in nontransformed and simian virus 40 (SV40)-transformed Balb/3T3 cells, in nontransformed and 3-methylcholanthrene (MCA)-transformed C3H/10T 1/2 cells, and in nontransformed and MCA-transformed AKR-2B cells. In contrast, increased plasma membrane phosphorylation was observed in MCA- and smoooth surface-transformed Balb/3T3 cells and in SV40-transformed Swiss 3T3 cells compared to the respective nontransformed parent cell lines. These observations suggest that increased cyclic AMP-independent phosphorylation of externally oriented plasma membrane proteins is not a universal marker for the transformed phenotype.

Effect of dithiothreitol on plasma membrane intramembranous particle topography in BALB/c 3T3 and simian virus-transformed 3T3 cells and plasma membrane vesicles.

A statistical analysis of plasma membrane intramembranous particle topography was performed on purified 3T3 and simian virus (SV)-transformed 3T3 plasma membrane vesicles and on intact 3T3 and SV3T3 cells. The results show that intramembranous particles were more aggregated in intact 3T3 cells than in intact SV3T3 cells. By contrast, in purified plasma membrane vesicles intramembranous particles were significantly more aggregated in SV3T3 than in 3T3 preparations. These unexpected results suggest that intramembranous particle topography is selectively altered in SV3T3 plasma membranes during the membrane vesicle isolation procedure. Inasmuch as the disulfide-reducing agent dithothreitol (DTT) was used to isolate membranes, tests were done to determine if DTT would preferentially affect particle topography in intact SV3T3 cells. Exposure of intact SV3T3 cells to 10 or 50 mM DTT induced intramembranous particle aggregation but had no effect on intact 3T3 cells. Preliminary studies also showed that other sulfhydrylreactive compounds preferentially induced particle aggregation in SV3T3 specimens. These observations suggest that the state of cross-linking of membrane-associated proteins by disulfide bonding influences intramembranous particle topography and that differences exist in plasma membrane-associated sulfhydryl proteins in nontransformed and transformed cells.

PRomotion of smooth surface tumorigenicity by phorbol myristate acetate in Balb/3T3 cells and Balb/3T3 T proadipocytes.

In vitro exposure of Balb/3T3 cells to phorbol and phorbol myristate acetate significantly increased their tumorigenic potential when implanted sc on smooth surface plates into syngeneic mice. This finding supports the hypothesis that many so-called normal cell lines may actually represent initiated cells that can be induced to become tumorigenic following exposure to promoting agents. Since the tumorigenic potential of many tissues in vivo is inversely proportional to the state of differentiation of the stem cell populations undergoing transformation, we assayed the relative tumorigenicity of Balb/3T3 T proadipocytes, which can differentiate in culture, and Balb/3T3 cells, which cannot differentiate in culture. The effects of tumor-promoting agents on these cells were also tested. Plate-implanted Balb/3T3 T proadipocytes were markedly less tumorigenic than Balb/3T3 cells, and Balb/3T3 T proadipocytes were not sensitive to the promotional effects of phorbol myristate acetate.

Ability of normal human keratinocytes that grow in culture in serum-free medium to be derived from suprabasal cells.

The current studies were performed to determine from which regions of the skin keratinocytes that grow in vitro are derived. Normal human foreskin specimens were first separated by differential trypsinization into two suprabasal fractions and one basal fraction. Utilizing complete MCDB 153 basal nutrient culture medium containing epidermal growth factor and insulin, we then evaluated the clonogenic potential of cells in these three fractions. Suprabasal cell fractions demonstrated a colony-forming efficiency as great as or greater than that of the basal cell fraction, and 10%-15% of the keratinocytes that grew in primary and secondary cultures expressed involucrin, a suprabasal keratinocyte differentiation marker. Of such involucrin-containing keratinocytes, 80% also possessed the potential to undergo DNA synthesis, as determined by autoradiography following a 48-hour incubation with [3H]thymidine. These observations support the conclusion that the human keratinocytes that grow in vitro in serum-free medium can be derived from suprabasal cells and, therefore, that a state of nonterminal keratinocyte differentiation exists.

Defective control of terminal differentiation and its role in carcinogenesis in the 3T3 T proadipocyte stem cell line.

The expression of defects in the control of terminal cellular differentiation has been implicated to be of etiological significance in the pathogenesis of cancer. However, it has not been established whether or not additional defects in the control of cellular differentiation and proliferation are required for the expression of the completely transformed phenotype. We therefore developed four clones of proadipocyte stem cells that express a single specific defect that limits their capacity to undergo the terminal phase in differentiation. The value of these clones is emphasized by the fact that they show no defects in their ability to regulate nonterminal differentiation or proliferation relative to native nontransformed proadipocyte stem cells. Terminal differentiation-defective clones were therefore assayed to determine if they express the completely transformed phenotype. The results show that differentiation-defective stem cells are not tumorigenic in vivo and do not grow in soft agar, which is an in vitro assay for expression of the transformed phenotype by murine mesenchymal cells. These data are interpreted to support the conclusion that the expression of defects in the control of the terminal phase of differentiation per se is not adequate to induce complete neoplastic transformation.

Defective cyclic adenosine 3'-5'-monophosphate-dependent phosphorylation of plasma membrane proteins in chemically and virally transformed cells.

Cyclic adenosine 3':5'-monophosphate (cyclic AMP)-dependent phosphorylation of endogenous plasma membrane proteins catalyzed by an endogenous plasma membrane protein kinase was assayed in purified plasma membrane preparations derived from nontransformed, methylcholanthrene-transformed, and simian virus 40 (SV40)-transformed BALB/3T3 cells. In nontransformed cells, cyclic AMP stimulated the phosphorylation of two proteins with molecular weights of 24,000 and 14,000. The labeling of these proteins could be inhibited by rabbit skeletal muscle protein kinase inhibitor. In methylcholanthrene-transformed cells, no cyclic AMP-dependent phosphorylation of endogenous plasma membrane proteins was observed. SV40-transformed cells also showed markedly decreased cyclic AMP-dependent phosphorylation of both endogenous plasma membrane substrates. Addition of exogenous cyclic AMP-dependent protein kinase from bovine kidney to plasma membrane preparations isolated from methylcholanthrene or SV40-transformed isolated from methylcholanthrene or SV40-transformed cells, however, catalyzed the cyclic AMP-dependent phosphorylation of both the M.W. 24,000 and M.W. 14,000 substrates. These data show that the plasma membranes of transformed cells have a defect in an endogenous cyclic AMP-dependent phosphorylation system and that this defect can be corrected by addition of an exogenous cyclic AMP-dependent protein kinase.

Resistance to neoplastic transformation induced by nonterminal differentiation.

Cell clones derived from undifferentiated 3T3 T mesenchymal stem cells show a low rate of spontaneous transformation but can be efficiently transformed by a variety of carcinogens. In contrast, it is now reported that cell clones derived from nonterminally differentiated 3T3 T stem cells are highly resistant to neoplastic transformation induced by physical and chemical carcinogens. Differentiation-induced resistance to neoplastic transformation is evident in both in vitro and in vivo transformation assays and can be stably expressed for greater than 50 population doublings. These results establish that resistance to neoplastic transformation can be regulated in mammalian stem cells by the process of nonterminal differentiation.

Independent alterations in cell shape and intramembranous particle topography induced by cytochalasin B and colchicine in normal and transformed cells.

Native differences in cell shape and plasma membrane organization in contact-inhibited and transformed cells and the effects of cytochalasin B and colchicine on these cells have been examined by scanning electron microscopy and freeze fracture-electron microscopy. Confluent BALB/c 3T3 cells show a flat, polygonal shape with limited cell overlapping, some microvilli, and plasma membranes with an aggregated distribution of intramembranous particles. Simian virus 40-transformed BALB/c 3T3 cells, by contrast, have a pleomorphic, bipolar spindle shape, extensive cell overlapping, more numerous surface projections, and a random distribution of intramembranous particles. Treatment of 3T3 and SV3T3 cells with 10(-6) M colchicine produced changes in cell shape and induced intramembranous particle aggregation in SV3T3 cells but did not significantly affect the freeze fracture morphology of 3T3 plasma membranes. Treatment of 3T3 and SV3T3 cells with cytochalasin B (1 mug/ml) also produced marked changes in cell shape and induced intramembranous particle disaggregation in 3T3 cells, but it did not affect intramembranous particle distribution in SV3T3 cells. Lower doses of colchicine (10(-9) M) or cytochalasin B (1 to 50 ng) modulated intramembranous particle distribution in transformed and normal 3T3 cells, respectively, without seriously affecting cell shape. These results are interpreted to suggest that modulation of cell shape or cell surface topography and intramembranous particle distribution are separable phenomena.

Neoplastic transformation and defective control of cell proliferation and differentiation.

The control of proliferation of nontransformed 3T3 t-proadipocytes in vitro can be mediated at three states in the G1 phase of the cell cycle. These states are induced by the commitment of cells to differentiate (GD); by growth factor deprivation at low density or "contact inhibition" at high density (Gs); and by nutrient deprivation (GN). To determine if neoplastic transformation of proadipocytes is associated with a selective defect in one or more of these G1 growth arrest processes, we developed and studied eight cloned and several noncloned tumorigenic proadipocyte cell lines. We report that all transformed proadipocyte cell lines are tumorigenic and all lack the ability to arrest at GD and differentiate. By contrast, or approximately 90% of transformed proadipocyte cell lines retain their ability to growth arrest at Gs at low density when deprived of growth factors, and or approximately 90% growth arrest at GN when deprived of nutrients. These observations suggest that neoplastic transformation of proadipocytes is primarily associated with abrogation of growth control mediated at GD. However, whereas most transformed proadipocytes arrest at Gs at low density when deprived of serum, all transformed proadipocyte cell lines do not efficiently arrest at Gs at high density due to "contact inhibition." This suggests that neoplastic transformation of proadipocytes results from a primary defect in growth control mediated at GD and from an additional defect at Gs. These results are discussed with respect to their possible significance for the biological mechanisms of the initiation and promotion of carcinogenesis.

Differentiation of fibroblast-like cells into macrophages.

Differentiated cells with the morphological, enzymatic, antigenic, and functional characteristics of macrophages formed when a variety of nontransformed and transformed fibroblast-like mouse embryo cell lines were grown in a medium supplemented only with human plasma. Differentiated cells contained numerous lysosomes and phagosomes, nonspecific esterase and acid phosphatase activities, and cell surface la antigens and were capable of phagocytosis of iron particles. Differentiated cells were also growth arrested in the G1 phase of the cell cycle, but both growth arrest and differentiation were reversible processes. These observations suggest that cells with the morphology of fibroblasts have the capacity to undergo nonterminal differentiation into macrophages.