Retinal pigment epithelium (RPE) degeneration is the hallmark of age-related macular degeneration (AMD). AMD, as one of the most common causes of irreversible visual impairment worldwide, remains in need of an appropriate approach to restore retinal function. Wet AMD, which is characterized by neovascular formation, can be stabilized by currently available therapies, including laser photocoagulation, photodynamic therapy, and intraocular injections of anti-VEFG  therapy or a combination of these modalities. Unlike wet AMD, there is no effective therapy for progressive dry (non-neovascular) AMD. However, stem cell-based therapies, a part of regenerative medicine, have shown promising results for retinal degenerative diseases such as AMD. The goal of RPE cell therapy is to return the normal structure and function of the retina by re-establishing its interaction with photoreceptors, which is essential to vision. Considering the limited source of naturally occurring RPE cells, recent progress in stem cell research has allowed the generation of RPE cells from human pluripotent cells, both embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSC). Since iPSCs face neither ethical arguments nor significant immunological considerations when compared to ESCs, they open a new horizon for cell therapy of AMD. The current study aims to discuss AMD, review the protocols for making human iPSCs-derived RPEs, and summarize recent developments in the field of iPSC-derived RPEs cell therapy. Age-related macular degeneration (AMD), the main cause of central vision loss in patients older than 55Y worldwide, is initiated by the degeneration and loss of the retinal pigmented epithelium (RPE) in the macula caused by diverse mechanisms that remain under investigation , 2. AMD Technologies have made clinical-grade cell replacement therapies from pluripotent stem cells (both ESCs and iPSCs) possible for AMD. Stem cells can differentiate into bonafide-like RPE cells in vitro, albeit the derivation of RPE from iPSCs is a much faster and more cost-effective approach , 12. OneAge-related macular degeneration (AMD) is the primary cause of permanent central visual loss globally. Clear central vision is needed for daily activities such as driving and reading. “Age-related” means that it occurs in older persons and “macular” comes from “macula,” which is responsible for sharp and high-accuracy vision in the central portion of the visual field . As longAs the world’s population ages, the incidence rate of AMD will increase significantly, and it has been estimated that it will affect around 288 million people by 2040. AMD is most prevalent in white patients, followed by Asians and Hispanics, and is lowest in the black ethnic group . Visual CHF) gene, and mitochondrial dysfunction [AMD physiopathology is not yet completely understood. Research has focused on a variety of mechanisms, including oxidative stress, chronic inflammation, complement cascade, single-nucleotide polymorphism in the complement factor H , age-related maculopathy susceptibility gene2 (ARMS2), and tissue inhibitor of metalloproteinase 3 (TIMP3) [CHF, which inhibits the inflammatory cascade that regulates inflammation [TIMP3 is strongly related to AMD development [TIMP3 modulates not only the action of MMPs but also other molecules, such as VEGF, EGF , and TNF (tumor necrosis factor), and thus, it has a fundamental role in maintaining the homeostasis of RPE extracellular matrix and RPE metabolism in AMD progression [The etiology of AMD may be attributed to genetic-related influences . Several (TIMP3) . The mosammation , 14. Theelopment . It has gression .trans-retinal into 11-cis-retinal, and finally, keeping the integrity of the RPE-retina structure by directional secretion of its necessary factors [The pathology hallmark of AMD is RPE–Bruch’s membrane complex damage. The RPE is a post-mitotic single sheet of cells  lying at factors . With ag factors .Fig. 1ScAs discussed previously, there are two main categories for AMD: dry or non-neovascular AMD and wet or neovascular AMD. Currently, no effective treatment is available for dry AMD . AlthougAvailable therapeutic options for wet AMD focus on limiting the neovascular membrane but do not repair the damage that may have already occurred. First-line therapy for patients who suffer from wet AMD is intravitreal VEGF inhibitors  .Photodynamic therapy with/without anti-VEGF medications is another option for the treatment of patients where initial treatment with anti-VEGF was not effective. Thermal laser photocoagulation can result in enlarging scotoma or a new scotoma development, so it is rarely recommended nowadays .Cell therapy offers an unlimited source of cells for cell transplantation studies . CurrentThe use of the ESCs technique has been associated with ethical limitations and immunological complications upon allogeneic transplantation  when theiPSCs are induced reprogramming of differentiated somatic cells back into an embryonic-like pluripotent status. iPSCs technology was established by Shinya Yamanaka, who showed that ectopic expression of four pluripotency transcription factors, termed KLF4, c-MYC, OCT4 and, SOX2, could convert somatic cells to the pluripotent state, which can then be re-differentiated into various desired types of cells . While iRetina has a complex architecture made up of the interconnection of a wide variety of cells . DegenerThe most important challenge facing cell therapists in treating AMD is choosing the source of cells and methods to generate bonafide RPE cells. However, since both hESCs and hiPSCs can differentiate toward RPE cells, controlling the potency of hPSCs differentiation into desired cells is one of the important goals of many research teams . AlthougVarious growth factors and chemical molecules have been tested on RPE production, including WNT antagonists (Dkk-1), bone morphogenetic protein (BMP) antagonist (noggin), activin A, antagonists of NODAL  ligand), insulin-like growth factor (IGF), and small chemical molecules, such as nicotinamide (vitamin B3), dorsomorphin, XAV939, SB431542, and heparin . The tim, Leach et al. published a reliable and rapid protocol for direct differentiation, in comparison with spontaneous differentiation protocols, that allow the efficient differentiation of RPE from iPSCs by combining factors such as activin A and nicotinamide (NIC) [Based on the investigation of developmental studies, scientists have designed different protocols with different combinations of cytokines and small chemical molecules . For insde (NIC) , in addide (NIC) , 54. Anode (NIC) .There is still controversy surrounding the immunogenicity of iPSCs and their derivatives, despite a report that differentiated cells from iPSCs are unlikely to be rejected by the immune system because they exhibit a limited immune response . AdditioSugita et al. (2020) developed an optimized good manufacturing practice (GMP)-compliant protocol for the conversion of iPSCs into RPE cells, in which human iPSCs were cultured on dishes coated with gelatin. In their protocol, signal inhibitors Y-27632, CKI-7, and SB43542 were added to the GMEM medium along with a knockout serum replacement. Following the observation of RPE-like colonies, they changed the medium to DMEM supplemented with B27 and L-glutamine. They also added SB431542 and FGF-2 to the culture .Zhu et al. utilized sequential retinal inducer factors  and RPE specification factors  to generate RPE from hiPSCs . Yang etIn a recent study, Michelet et al. introduced a simplified 2D culture in combination with lipoprotein uptake-based sorting  to derive an RPE monolayer from hiPSCs within 90 days. The author mentioned that differentiation of RPE by this protocol obviates the need for growth factors and small chemical molecules; thus, the production of RPE by this protocol is more cost-effective. A feeder-free culture system is also preferred .For clinical uses of hiPSCs, Takao Kuroda et al. preferred a feeder-free culture in their reports . They shIn another model of differentiation, Ye et al. found that sequential treatment with inhibitors of signaling pathways  plus nicotinamide can increase the purity and quality of RPE sheet generation. In their experiments, they did not use any artificial scaffolds for RPE sheet transplantation since artificial scaffolds may cause inflammation; also, they did not report any tumor formation and immune rejection after transplantation. As mentioned in the Ye et al. study, effectual production of pure RPE sheets combined with the assistance of a noninvasive model that used F-actin-labeled images for machine learning-based TER prediction will be valuable for quality control and large-scale manufacturing of RPE sheet for clinic, industry, and facilitation of cell therapies .OTX2, PAX6, and MITF could drive RPE differentiation in iPSCs. These transcription factors are critical regulators during eye development, the process by which anterior neuroectoderm cells become progressively specified to the RPE lineage. Overall, these transcription factors work together to promote RPE development by specifying and maintaining the eye field, the optic vesicle, and RPE [In reprogramming and differentiation  studies, robust protocols have been developed to drive iPSCs differentiation to specific types of cells through overexpression of specific cell lineages transcription factors . Inspire and RPE .According to selected references, Table There are two methods for RPE cell therapy of the retina: RPE cell suspension injection and the insertion of a monolayer patch of RPE sheet seeded on special scaffolds into the subretinal space . A simplTo date, the only hiPSC-derived RPE transplantation study was conducted on the implantation of scaffold-free RPE sheets to treat chronic wet AMD . In thatThe safety and the feasibility of RPE cell suspension injection and RPE patch implantation approaches were checked in phase I/II clinical trials and have shown promising results . For the. Despite progress in stem cell research, scientists are faced with different challenges such as ethical issues, regulatory controversies, safety, and efficacy, along with the technical difficulties of adjusting this method into a standard approach for clinical application. Before iPSCs can be considered a reliable cell source for clinical-grade purposes, a variety of concerns should be taken into consideration [According to several studies, the morphology and function of iPSC-derived RPE cells were similar to naïve RPE in vivo and in vitro . Despitederation . For insderation .iPSC-RPE transplantation is the most challenging procedure due to immune responses. Clinical trials have noted that RPE allografts failed to survive due to immune rejection . The rejMaking the target cells free of pluripotent stem cells is also a major challenge in cell therapy. Undifferentiated pluripotent stem cells are master cells that can create various cells of the three embryonic germ layers, and they can carry the risk of tumor formation. iPSCs were transplanted subcutaneously into immunosuppressed mice in a study to confirm their tumorigenicity. This study showed that transplanted iPSCs are tumorigenic and able to evade immune detection . Hence, Generic mutations in iPSCs or their derivation are of concern since they increase the risk of cancer development in patients and the risk of transformation of the cells. For instance, the initial Japanese study that used autologous iPSC-derived RPE cells for the treatment of AMD was halted because reprogrammed iPSCs from the second patient showed unexpected mutations . TherefoPrior to clinical-grade use of iPSC and its derivations, it is necessary to check for cross-contamination of the cell lines .Since stem cells cannot decontaminate themselves, their microbiological sterility is vital in order to prevent mycoplasma, bacterial, viral, and fungal contamination, which is evident in cell transplantation therapies . A complIt is crucial for clinical-grade iPSCs to have high efficacy during reprogramming. Studies demonstrate that small-molecule inhibitors  can increase the efficacy of the reprogramming procedure significantly .A multitude of therapeutic options based on stem cells has been explored over the last several decades. Since induced pluripotent stem cells (iPSCs) are less immunogenic and have less ethical controversy than hESC-based therapies, exploring their therapeutic potential is particularly intriguing. iPSC-derived RPE transplants became available novel treatment to humans after a decade of preclinical studies to restore vision for the patient who suffers from AMD. There has been enough evidence produced so far to confirm the safety of these potential therapeutic approaches in phase I/II clinical trials. Thus, there is probably less time to go until we have a stem cell-based treatment for acute wet AMD since only RPE cells with Bruch's membrane need replacing. However, we are far from being able to treat late dry AMD because the chronic loss of RPE will also result in secondary loss of photoreceptors overlying the affected retina. As discussed in this review, further studies must take advantage of the manufacturing process and subretinal delivery of the transplanted cell to improve the efficacy of RPE fabrication and their integration into the retina as well as improve the retina microenvironment for long-term integration and survival of transplanted cells.


Cao et al.). Cells, scaffolds and growth factors represent the key elements of tissue engineering. Due to the ability of the bone to self-regenerate due to the migration of cells and growth factors from the adjacent tissues, the success of bone tissue engineering largely relies in the ability of the scaffold to act as a template for guiding tissue regeneration. Indeed, not only cell-laden scaffolds, but also cell-free biomimetic matrixes are considered appealing solutions in bone regeneration, as they can stimulate cell colonization and recruitment from neighbouring tissues. In this context, additive manufacturing (AM) technologies have gained considerable interest owing to their versatility and the possibility to produce personalized scaffolds with complex geometry, matching the patient’s bone defects .In the attempt to repair bone tissue defects, a plethora of bone substitute materials of various origins have been utilized so far. Despite the fact that autogenous, allogenic and xenogeneic bone grafts still constitute valid therapeutic options in daily practice, bone tissue engineering strategies have gained popularity for the regeneration of damaged tissues . Nevertheless, the routine clinical use of AM is still very limited and is based on single case reports .Although several endeavours intended to promote vertical bone augmentation have been reported, they are characterized by inconsistent long-term results and varying degrees of success . AM is eLatimer et al.). As an alternative cell delivery method, cell-encapsulated hydrogel-scaffold constructs have been developed for the controlled release of stem cells at the surgical site. When 3D cell cultures are preferred over monolayer cultures, the encapsulation of the self-assembled cell aggregates within a hydrogel has the advantage to protect the 3D assemblage from disaggregation. In an ectopic bone formation model in mice, a 3D printed polymeric scaffold was found to promote ectopic mineralization to a higher extent when combined with a hydrogel containing spheroid bone marrow-derived stem cells (BMSCs), than when combined with a hydrogel laden with dissociated BMSCs .Combining stem cells with 3D scaffolds may represent a step forward in the pursuit of a faster and improved bone healing. Cells can be directly seeded onto 3D scaffolds before implantation or can be incorporated within biodegradable scaffolds during the manufacturing process using bioprinting techniques .Other strategies to foster bone tissue ingrowth and mineralization within porous additive manufactured scaffolds rely in the chemical and topographical modification of their surfaces. An acellular organic-inorganic mineralizing construct has been successfully produced, combining a 3D printed nylon scaffold with an elastin-like recombinamer coating, that can be pre-mineralized in the lab prior to implantation . As such, 3D collagen-based matrices functionalized with BMP-2 or with an enamel matrix derivative were found to stimulate in vitro the osteogenic commitment of osteoprogenitor cell lines .Scaffold bio-functionalization with bioactive agents offers the great advantage of inducing stem-cell homing, while avoiding the several concerns related to stem cell implantation. Among various molecules, the incorporation of bone morphogenetic protein 2 (BMP-2) in 3D scaffolds has been widely explored, demonstrating its ability to trigger stem cell osteogenic differentiation .Beside exogenous growth factors, autologous platelet concentrates (APCs) can be employed to enhance the biological properties of bone substitutes. Promising Latimer et al.). Up-to-date periodontal regeneration strategies include the use of stem cells and the design of multi-material and micropatterned scaffolds, favouring compartmentalized tissue healing and periodontal fiber orientation. Despite the significant advancements noticed in this field, further studies are required to improve the complex multi-tissue periodontal regeneration . Furthermore, different tissue engineering strategies, including and not limited to the use of bio-printing, have been introduced to regenerate dental tissues . However, the development of fully formed functional teeth seems far from being achievable in the near future. A deeper understanding of cell-cell and cell-matrix interactions during tooth formation, together with progresses in AM, is needed so that bio-engineered teeth become a clinical reality.In dentistry, bone augmentation procedures are performed in the vast majority of the cases to restore adequate alveolar ridge dimensions for dental implant placement . Beside Zhu et al.).This issue focused mainly in bone regeneration in the oral and maxilla-facial area. However, a broader view on recent developments in the area of orthopaedics was also maintained. Therefore, clinically challenging topics such as non-union fractures and delayed bone healing can also be addressed in the information provided in this issue .Taking into consideration the ageing population and the fact that chronic diseases associated with bone healing and metabolism  have high prevalence, novel ways addressing bone healing should be introduced. In this context, the discovery of selective drugs for targeting osteoporosis is particularly relevant. In this issue, an Wang et al.).However, bone fractures are not limited only to osteoporotic conditions. Special attention has to be devoted to the treatment of pathologies occurring at the growth plate in the paediatric age. This cartilaginous region, which acts as the primary centre for endochondral bone formation in immature long bones, is particularly susceptible to fractures. Current treatments often lead to the development of an undesired bone bridge and to related growth disturbance risks. AM, in combination with cell seeding and active substance delivery, could offer alternative options to existing clinical solutions, in order to achieve cartilaginous tissue reconstruction at the growth plate, thus avoiding the formation of calcified physeal scars (Overall, AM, in combination with tissue engineering strategies, offers emerging opportunities in bone regeneration, by enabling the production and the biofunctionalization of customized site-specific 3D scaffold with tunable properties. Looking at the future, remarkable efforts should be directed to the optimization of current technologies and to overcome the hurdles that are delaying the translation of addictive manufactured tissue-engineered bone scaffolds into everyday clinical use.


Engineered probiotics are a kind of new microorganisms produced by modifying original probiotics through gene editing. With the continuous development of tools and technology progresses, engineering renovation of probiotics are becoming more diverse and more feasible. In the past few years there have been some advances in the development of engineered probiotics that will benefit humankind. This review briefly introduces the theoretical basis of gene editing technology and focuses on some recent engineered probiotics researches, including inflammatory bowel disease, bacterial infection, tumor and metabolic diseases. It is hoped that it can provide help for the further development of genetically modified microorganisms, stimulate the potential of engineered probiotics to treat intractable diseases, and provide new ideas for the diagnosis of some diseases or some industrial production. Probiotics have been studied for decades. The word "probiotics" first appeared in 1974, and, in 2001, the Food and Agriculture Organization of the United Nations and the World Health Organization defined probiotics as "live microorganisms which when administered in adequate amounts confer a health benefit on the host" . ProbiotIn recent years, genome sequencing has become more affordable and some of the tools for editing and modifying microbial genomes have become more powerful, enable us to engineer probiotics according to our own ideas, so that we can develop customized probiotics . By meanThrough gene editing, the existing probiotics are modified to obtain the desired new probiotics. Such engineering modification allow us to directly verify whether the genetic material, proteins and functional roles of these novel microorganisms have been changed as desired. Gene editing technology continues to develop, from homologous recombination to the first-generation Zincfinger Nucleases (ZFNs) technology, to the second-generation Transcription Activator-like Effector Nucleases (TALEN) technology and to the more popular third-generation Clustered Regularly Spaced Short Palindromic Repeats CRISPR associated (CRISPR-Cas) technology in recent years . Among tThese gene-editing tools introduce Double Strand Break (DSB) into target genes, DNA Repair is induced by an error-prone Non-homologous End Joining (NHEJ) pathway or Homology Directed Repair (HDR) . ZFNs anStreptococcus thermophilus [CRISPR and related Cas genes are considered to be part of the bacterial immune system, which is a self-protective sequence of bacteria and can provide phage resistance . At the mophilus , It snipmophilus . By artimophilus .CRISPR technology can be used not only for the study of loss of function such as CRISPR knock out (CRISPRKO) and CRISPR interference or inhibition (CRISPRi), but also for the screening study of gain of function such as CRISPR activation (CRISPRa) . The devAdvances in gene editing technology provide new possibilities for gene editing of probiotics. Engineered probiotics have developed rapidly in recent years. Researchers have conducted a variety of biomedical studies on engineered probiotics for disease diagnosis and treatment. In particular, several engineered probiotics have also entered the clinical trial stage Table . ScientiMicrobiota imbalance is closely related to the development of inflammatory bowel disease (IBD). Probiotics have been shown to improve the microbiome of the microbiota by changing the intestinal environment and inhibiting the growth of harmful bacteria, also, they can prevent inflammatory diseases from further affecting the host immune system, and have positive significance for the regulation of inflammation , 46. AdvE. coli Nissle 1917 (ECN) to produce a curly fiber matrix that promotes intestinal epithelial integrity, and the trefoil factor (TFF) component in the matrix promotes intestinal barrier function and epithelial repair [E. coli Nissle 1917 to inhibit the excessive immune response in the local tissues of mouse colitis and regulate the intestinal microbiota to promote the reversal of inflammation [Specific probiotics have been designed to diagnose and treat IBD. Around 2017, researchers have used engineered bacteria to diagnose IBD –17. Basel repair . Their sl repair , 19, 28.ammation . Liu et Bacterial infections have a great impact on people’s health and nearly one million people die from bacterial infections worldwide each year, and the lack of bacterial resistance and new antibiotics has become an obstacle to solving the problem of bacterial infections . ResearcLactobacillus casei strain that can produce Listeria adhesion protein [Listeria mucosa and systemic transmission, protecting mice from fatal infections. They can also enhance the intestinal immune regulation function by accumulating intestinal mucosal regulatory T cells, CD11c+ dendritic cells and natural killer cells. The method of engineering L. casei ATCC334 to reduce Listeria infection and protect the intestinal tract is worth learning. Some researchers have designed and constructed an engineered probiotic based on E. coli Nissle 1917, which can specifically target and kill the two most common vancomycin-resistant Enterococcus, and significantly reduce the number of Enterococcus faecalis and Enterococcus faecium in the feces of model mice [Saccharomyces boulardii so that the engineered bacteria secrete a fusion protein (ABAB) that can neutralize 4 different Clostridium difficile toxins [Clostridium difficile in the mouse intestines mucosa, therefore reduce the mortality of these mice. In addition, researchers have developed some engineered probiotics to better prevent and prevent and diagnose bacterial infections. Some researchers have genetically modified Lactobacillus lactis to produce engineered probiotics that recognize the cholera autoinducer 1 (CAI-1) produced by Vibrio cholerae and cause color changes in feces [lactic acid bacteria that can be used to detect real-time changes in autoinducer peptide-I (AIP-I) produced by Staphylococcus aureus [. The researchers also engineered E. coli Nissle 1917 to specifically kill pathogens by detecting the autoinducer N-acyl homoserine lactone (AHL) of Pseudomonas aeruginosa and releasing antibiotics and anti-biofilm enzymes [Some researchers have successfully developed engineered probiotics. The American team of Arun K. Bhunia designed an engineered  protein . This stdel mice . Some ree toxins . The pres aureus . The res enzymes . This seThe discussion about the treatment of cancer by bacteria has existed for decades , 49. SomE. coli strain subtype called "SLIC" [E. coli Nissle 1917 to target the angiogenic inhibitor TUM-5 and tumor suppressor p53 to the anaerobic tumor region [E. coli Nissle 1917 that can be targeted to colonize tumors has also been published recently. It can convert the metabolic waste produced by tumors into L-arginine that enhances the anti-tumor immune response, and can effectively enhance the therapeutic effect of programmed cell death protein-1 and its ligand inhibitors on mouse tumors [E. coli Nissle 1917 that can achieve precise tumor treatment in vivo through the control of light color changes has also been successfully constructed(9). These studies have provided some help for the treatment of tumors. Now there are two engineering probiotics related to the treatment of tumors in clinical trials (NCT04167137 and NCT04025307). The most notable of these is the SYNB1891 strain obtained by the engineering of E. coli Nissle 1917 by Synlogic (NCT04167137). This strain can produce cyclic di-AMP to stimulate the interferon gene pathway, and then trigger innate immunity by activating antigen-presenting cells to present tumor antigens [In the study of Candice R. Gurbatri and others at Columbia University, researchers used synthetic biology methods to modify bacteria and obtained an engineered d "SLIC" . SLIC coe tumors . An engiantigens . Their pThe content of metabolism-related substances is generally in a relatively stable state in the body. When the biochemical process in the body is hindered, the accumulation or lack of certain metabolites can cause diseases –55. ThesLactobacillus reuteri 100-23C, so that the engineered probiotic strain can produce phenylalanine lyase, which can reduce the blood phenylalanine of the PKU mouse model [E. coli Nissle 1917, and constructed a strain of E. coli named SYNB1618 [Phenylketonuria (PKU) is caused by mutations in the phenylalanine hydroxylase gene. Patients with phenylalanine hydroxylase deficiency cannot effectively decompose phenylalanine, causing phenylalanine to accumulate in the blood, which seriously affects the patients’ health . The resse model . Their eSYNB1618 . This enSYNB1618 . This faE. coli strain called SYNB8802, which can ingest oxalic acid in the gastrointestinal tract to reduce urinary oxalic acid levels and reduce kidney damage caused by hyperoxaluria. Results from a Phase 1a study of this trial demonstrated the strain's safety profile, and the later trials are still in progress (NCT04629170). Novome has also developed a drug named NOV-001 for the treatment of intestinal hyperoxalic acid. This drug is composed of engineered strain NB1000S and plant-derived polysaccharide NB2000P. It is now undergoing clinical trials to verify its efficacy (NCT04909723). In our opinion, engineered probiotics could be a good way to treat diseases in combination with other drugs.Mutations in the alanine glyoxylate aminotransferase gene can cause hyperoxalic acid. The accumulation of glyoxylic acid can lead to the increase of oxalate and even the production of urinary stones . SynlogiE. coli Nissle1917 to obtain an engineered strain named SYNB1020. This strain can convert ammonia into L-arginine, reduce the ammonia content in the intestine of a mouse model of hyperammonemia, and improve the survival rate of mice [Ammonia is a highly neurotoxic metabolite. Impaired ammonia clearance occurs when urea cycle enzyme deficiency or liver cell dysfunction occur. If hyperammonemia is not treated in time, coma or even death may occur . Researc of mice . HoweverLactobacillus gasseri that secretes glucagon-like peptide-1 (GLP-1) to improve diabetes [Lactococcus lactis to obtain a engineered bacteria named AG019. AG019 can deliver proinsulin and interleukin-10 to the gastrointestinal mucosa tissues, reduce or eliminate the damage to pancreatic β cells, and potentially stabilize or improve endogenous insulin production. Their experiment Also in progress (NCT03751007).Researchers have also developed engineered probiotics for some other metabolic-related diseases. Some researchers have constructed an engineered commensal diabetes . GLP-1 sdiabetes . PrecigeLeuconostoc citrate [E. coli LS31T to increase the production of riboflavin [E. coli that can further convert the products of photosynthetic cyanobacteria into 2,3-butanediol [E. coli strain that can produce benzoic acid from plant-derived glucose [Clostridium ljungdahlii to produce isopropanol, 3-hydroxybutyric acid, ethanol and other substances [E. coli, lactic acid bacteria and yeast into engineered bacteria [Engineered probiotics can not only be used for the diagnosis and treatment of some diseases, but also be closely related to industrial production. Jaewoo Son and others of the Korea Academy of Science and Technology used the CRISPRi system to develop an engineered  citrate . The strboflavin . Thus thtanediol . The stu glucose . Their rbstances , which rbacteria –66.Bacillus subtilis, which can further convert the acetaldehyde converted from ethanol into acetic acid to reduce the harm of alcohol. They used the spores of this strain to conduct a 90-day repeated administration toxicological evaluation on mice, and proved that the bacterium has no side effects [Lactobacillus plantarum WCFS1 to produce n-acetylglucosamine [Brucella ATCC MYA-796 can ferment galactose faster [Corynebacterium glutamicum, Candida parapsilosis, Lactobacillus casei, Lactobacillus reuteri, Bifidobacterium, Clostridium butyricum, and Escherichia coli [Zbiotics has added an acetaldehyde dehydrogenase gene that decomposes acetaldehyde in  effects . Zbioticcosamine . Some ree faster , 70. Thehia coli –78. TheyThe emerging gene-editing methods used to develop synthetic engineered probiotics have great potential for the treatment and diagnosis of different types of human diseases, and engineered probiotics may become a new method for the treatment of cancer, inflammation, infection and some other diseases. The engineered probiotics designed by the researchers are more effective, have fewer side effects and are more affordable than traditional treatments or wild-type strains, benefiting more patients and families. The advantages of engineering probiotics such as stability, specificity, preference, low cost, and relative safety may make them a new choice for the treatment of many of the diseases mentioned above and more. However, the application of engineered probiotics still faces a series of challenges. For example, many microorganisms have few genetic tools that can be manipulated, and the microorganisms currently used for research design are only a small part of the vast family of microorganisms, which leads to the restriction of probiotic modification to a few strains. There are still a few engineered probiotics that go from the laboratory to the clinic, and even if they enter clinical trials, it cannot be guaranteed that it can achieve the intended effect, which hinders the efforts of some researchers to construct engineered probiotics. Some argue that genetically modified organisms (GMOs) are harmful, preventing the effective deployment of engineered probiotic therapies.Despite these limitations in the research and application of engineered probiotics, we cannot ignore their potential role in improving human health. Success and failure are inevitable in the research and development of engineered probiotics. We need more research to find the possibility of success, not only in the research of various probiotic modification tools, but also in-depth understanding of disease pathology, so as to make it easier for us to design the desired probiotics. The efficacy of engineered probiotics is a problem that we must consider, but before these probiotics undergo clinical transformation, safety issues must be resolved. It is necessary to ensure that the guidelines established by various health regulatory agencies are followed to design and construct safe and effective probiotics. We believe that with the increasing understanding of the human microbiome and disease mechanisms, the rapid development of gene editing technology, safe, stable and effective engineered probiotics will surely shine in many fields.


This Research Topic focused on illuminating the role of natural products in amelioration of oxidative stress, inflammation, and cell death in metabolic and cardiovascular diseases. Although these processes are essential in maintaining homeostatic cell signalling functions under physiological conditions; however, their exacerbation disturbs tissue homeostasis and promotes cell death. The common link for long-term cardiovascular pathologies is oxidative stress, resulted due to an imbalance between the generation of free radical oxygen species (ROS) and endogenous antioxidant enzyme activity. Oxidative stress, inflammation and cell death are implicated in the etiology or pathology of cardiovascular disorders such as chronic heart failure, angiogenesis, atherosclerosis, myocardial infarction, and pulmonary hypertension, accountable for an increase in stroke incidence amongst young populations . AccordiIn recent times, several studies have emphasized the importance of bioactive natural compounds for the prevention and treatment of diabetes and cardiovascular diseases. Medicinal plants or natural products are valuable sources of antioxidants that can help fight against detrimental oxidative stress, inflammation, and cell death, which are exacerbated during cardiovascular disease progression. Natural products therapy for the treatment of metabolic and cardiovascular diseases has shown encouraging results, and declined the mortality rate of these patient populations. Although the progress of natural product research on cardiovascular diseases is satisfactory, a huge number of these natural products remain unexplored, and it warrants a more comprehensive understanding of their beneficial effects. The current Research Topic was conducted to explore recent advances in natural product treatment for cardiovascular diseases, in order to provide valuable clues for promising therapeutic targets and new pathological mechanisms. The Research Topic published seven articles, of which two are review articles and five are original contributions. These articles shed a light on the importance of treatment with natural compounds for the mitigation of cardiovascular disorders. The main goal of this editorial is to provide the roadmap for the quality publications that may update the knowledge about the beneficial effect of natural products for the treatment of cardiovascular diseases.An et al. emphasized the significance of the antioxidant effects of berberine in preventing cardiovascular disease progression in preclinical and clinical settings. They comprehensively discussed the mechanism of action of berberine in preventing cardiac oxidative stress, providing novel insights into berberine usage in the management of heart failure, arrhythmia, myocardial ischemia/reperfusion, and coronary atherosclerosis. Angiogenesis, the formation of new blood vessels from pre-existing blood vessels, plays a crucial part in the amelioration of cardiovascular pathologies, such as ischemic cardiac diseases and stroke. Li et al. comprehensively reviewed the mechanism of action and potential applications of several pro-angiogenic phytoconstituents including ginsenosides from Panax notoginseng, astrgalosides, and calycosin from Radix Astragali, paeoniflorin from Radix Paeoniae, salvianolic acid B from Salvia miltiorrhiza, ferulic acid from Angelica sinensis, ilexasponin A1 from Ilex pubescens, and puerarin from Radix puerariae in ischemic heart diseases. They highlighted that these pro-angiogenic phytochemicals ameliorate myocardial ischemic diseases by reducing oxidative stress, inflammation, and cell death. These outcomes aid in the identification of drug candidates from medicinal plants for the treatment of atherosclerotic and ischemic cardiovascular diseases.In this Research Topic, you would find two interesting reviews about the potential applications of berberine and herbal medicine in the treatment of cardiovascular diseases. Li et al. investigated the importance of Qishen granule (QSG), a clinically approved traditional Chinese medicine in attenuating cardiac inflammation. The authors employed left anterior descending ligation and splenectomy to explore the QSG effect on inflammation induced by the cardio-splenic axis, which dominates the inflammation injury during heart failure. QSG treatment improved cardiac function, inhibited the release of splenic monocytes, and reduced macrophage recruitment and activation in the acute myocardial infarction-induced heart failure mouse model. Furthermore, QSG downregulated Toll-like receptor 4 (TLR4)/MyD88/nuclear factor (NF)-κB p65 inflammatory pathway to counteract cardio-splenic monocyte activation. Therefore, QSG could be a promising anti-inflammatory therapy for the treatment of heart failure and delay the progression of heart failure.To provide new insights for the treatment of heart failure, the molecular signalling mechanisms of different medicinal plants or natural products, are included in this Research Topic. Heart failure is the most debilitating public health concern with high rate of mortality and morbidity globally. In chronic heart failure, oxidative stress and inflammation occur in the myocardium and promote hypertrophic signalling, cardiac cell death, and cause left ventricular dysfunction and arrhythmia . To counZhang et al. revealed the therapeutic mechanism of the Shexiang Tongxin Dropping Pill (STDP), a traditional Chinese Medicine for chronic heart failure. STDP treatment relieved cardiomyocyte hypertrophy, myocardial fibrosis and improved cardiac function in transverse aortic constricted (TAC) mouse, a chronic heart failure mouse model. Using network pharmacology analysis and whole-transcriptome sequencing, the authors revealed that STDP is a “multi-component, multi-target” drug, and shows great potential for the mitigation of chronic heart failure. Interestingly, STDP treatment normalized the 395 imbalanced genes of chronic heart failure in the TAC sugery mouse model. Of note, STDP targets signalling pathways related to fibrosis, energy metabolism, and inflammation by inhibiting transforming growth factor (TGF-β) and ERK/mitogen activated protein kinase (MAPK) pathways. In a similar manner, using the network pharmacology approach, Guo et al. revealed that the calcium signalling pathway is a molecular target for the Danhong injection (DHI) in the prevention of acute myocardial infarction. DHI regulates the calcium signalling pathway, of importance, by inhibiting phospholamban (PLB), sarcoplasmic reticulum Ca2+ ATPase (SERCA), and calcium/calmodulin-dependent protein kinase II gamma (CaMK II) protein expression to improve acute myocardial infarction. Additionally, DHI is effective in reducing the key pathological changes of acute myocardial infarction such as the inflammatory cell intrusion, myocardial cell edema, and degeneration in the mouse models. These studies suggest that STDP and DHI are new treatment options for the prevention and treatment of heart failure, which can interfere with the process of cardiovascular diseases from different aspects.In another study, Wang et al. demonstrated that Dan-Shen-Yin (DSY) granules are effective in ameliorating hypoxia-induced pulmonary hypertension (HPH). 147 potential targets of DSY on HPH were identified based on network pharmacology, of which, oxidative stress , inflammation , cell proliferation , and PI3K-Akt signalling pathway are highly enriched by DSY. In addition, authors demonstrated that DSY targets AKT serine/threonine kinase 1 (AKT1), signal transducer and activator of transcription 3 (STAT3), and hypoxia-inducible factor-1 (HIF-1) proteins to decrease the detrimental effects of HPH, using a hypoxia-induced mouse model. They observed that DSY therapy downregulated proliferating cell nuclear antigen (PCNA) in lung tissues through suppression of the STAT3/HIF-1α and FAK-AKT signalling pathway. Therefore, we can conclude that DSY can prevent the development of HPH and alleviate pulmonary vascular remodelling through multiple pathways.Pulmonary hypertension is a complex and serious condition characterized by high blood pressure in the lung arteries. In a chronic cardiopulmonary dysfunctional state, progressive remodelling of pulmonary arteries and the building up of elevated lung arterial resistance lead to right ventricular failure and death . There aHosseini et al. evaluated the protective effect of Rheum turkestanicum on doxorubicin-induced cardiotoxicity due to its substantial in vitro antioxidant activity. The central outcome of this study is that Rheum turkestanicum inhibited doxorubicin-induced cardiotoxicity and decreased pathological abnormalities by controlling oxidative stress and antioxidant defenses in the cardiac tissue. The authors suggested that Rheum turkestanicum might be beneficial as a complementary strategy for reducing digoxin-induced cardiotoxicity.According to these studies, it is hypothesized that reducing oxidative stress, inflammation, and cell death is beneficial in treating cardiovascular disorders such as heart failure and pulmonary hypertension. The current findings are impressive, but further research is needed before natural products may be used to treat metabolic and cardiovascular problems. First, it is very difficult to determine their clinical usefulness due to a scarcity of clinical validation examining these natural compounds’ long-term impact on cardiovascular outcomes. Second, a contentious issue that undermines the favorable therapeutic effects against cardiovascular illnesses is the limited systemic bioavailability of phytochemicals and nutraceuticals. Is it still unclear whether novel drug delivery techniques will increase the systemic bioavailability of these agents? Furthermore investigating any possible pharmacokinetic or pharmacodynamic interactions between natural products and medications provided for CV patients is important for its integral therapy. Natural products should be taken into account as a crucial component of therapy despite these limitations, especially for patients who fall into the high and very high-risk categories because they may assist to reduce residual risk and achieve treatment objectives. We think that the most recent research on a variety of natural products that are highly relevant to the treatment of cardiovascular disorders was published in this Research Topic.


The problem of drug resistance represents a major challenge for the cure of non-small cell lung cancer. Tumor cells can be intrinsically resistant to drugs or they can become resistant during the treatment as a result of several adaptive responses. However, molecular mechanisms at the basis of drug resistance are not fully understood. MicroRNAs are small non-coding RNA that regulate gene expression and play critical functions in many cellular processes. Recently, the study of microRNAs has provided evidence about their role in the regulation of molecular mechanisms at the basis of drug resistance. Here we summarize the available knowledge about the role of miRNAs in the resistance to drugs that are currently used to treat non-small cell lung cancer and we critically discuss the experimental approaches on the basis of this evidence.Lung cancer is the leading cause of cancer-related mortality in the world. The development of drug resistance represents a major challenge for the clinical management of patients. In the last years, microRNAs have emerged as critical modulators of anticancer therapy response. Here, we make a critical appraisal of the literature available on the role of miRNAs in the regulation of drug resistance in non-small cell lung cancer (NSCLC). We performed a comprehensive annotation of miRNAs expression profiles in chemoresistant versus sensitive NSCLC, of the drug resistance mechanisms tuned up by miRNAs, and of the relative experimental evidence in support of these. Furthermore, we described the pros and cons of experimental approaches used to investigate miRNAs in the context of therapeutic resistance, to highlight potential limitations which should be overcome to translate experimental evidence into practice ultimately improving NSCLC therapy. Lung cancer represents the main cause of cancer-related death worldwide. Despite recent advances in lung cancer management, the prognosis of patients remains adverse with five-year survival rates ranging from 6% in the case of metastatic cancer  up to 33–60% in patients with localized tumours . Non-smaMiRNAs are non-coding RNAs (ncRNAs) of ~22 nucleotides, which act mainly as post-transcriptional gene regulators . The impSuch molecular and biological features of miRNAs make them play key roles in the regulation of physiological and pathogenic cellular processes relevant to cancer progression and therapy response.Here, we reviewed the most relevant literature focused on the role of miRNAs and their molecular mechanisms at the basis of drug resistance in NSCLC. The main experimental strategies as well as their pros and cons were also discussed.We initially performed a literature search (see Methods) and reviewed a total of 502 publications (from 2009 to June 2022) that report signatures of miRNAs involved in the responses to different drugs used to treat NSCLC patients . Next, wOut of a total of 42 drugs used in the NSCLC clinical setting , we foun−Cisplatin import/export: regulation of cisplatin import/export is an important mechanism of resistance to cisplatin. MiR-369-3p overexpression promoted cisplatin resistance by direct regulation of the expression of the SLC35F5 gene, a nucleotide sugar transporter involved in drug uptake . In addi−DNA damage repair: upon entering the cells, cisplatin becomes activated by the displacement of chloride atoms by water molecules. Active cisplatin exerts its cytotoxic function mainly by generating monoadducts and crosslinks at the level of the DNA. Therefore, genetic and epigenetic alterations of genes involved in DNA damage response represent a major mechanism to cope with cisplatin-induced cell death. In line with this, miR-92a-3p regulates cisplatin resistance by directly binding to the 3′ untranslated region (UTR) of RAD21 mRNA, a member of the cohesin complex that can promote DNA repair at the G2 phase of the cell cycle . Yet, mi−miRNA sponges: long non-coding RNAs (lncRNAs) are transcripts longer than 200 nucleotides that do not code for functional proteins. Since lncRNAs can contain MREs in their sequence, they can sequester miRNAs and impair their activity on target genes, therefore, acting as competitive endogenous RNAs or miRNA ‘sponges’. Over the last years, a high number of reports described lncRNAs that regulate cisplatin resistance by sponging specific miRNAs. For example, NORAD is a lncRNA upregulated by DNA damage and was found involved in cisplatin resistance by regulating SOX4 expression by sponging miR-129-1-3p . Similar−Pro-survival and apoptotic signaling pathways: when cells are exposed to platinum, both pro-survival and apoptotic signaling pathways are activated and compete for the final fate of the cells. A number of miRNAs have been associated with the direct or indirect regulation of apoptotic proteins or survival signaling pathways. MiR-103a-3p induces ERK signaling in NSCLC cells by targeting NF1 expression, a key negative regulator of the Ras signaling pathway . In an i−Reactive oxygen species: besides DNA damage, activated cisplatin is also a potent inducer of reactive oxygen species, which induces cell death. In this scenario, miR-495-3p overexpression was reported to modulate cisplatin resistance through direct inhibition of NRF2 , a trans−Epithelial-to-mesenchymal transition (EMT): EMT contributes to resistance to several therapeutic agents, including cisplatin . In line−Autophagy: autophagy is a crucial process that allows the recycling of important cellular components in response to stress conditions such as those induced by cisplatin treatment. Indeed, regulation of autophagy has been widely associated with cisplatin resistance phenotype. Rescue of miR-1-3p increases the sensitivity of cisplatin resistance cells by inhibiting ATG3, a key autophagic protein . FurtherOver the last 30 years of research, a variety of molecular mechanisms were described to be involved in the onset of resistance to cisplatin which were extensively reviewed elsewhere . Yet, a + T cells tumor infiltration, inducing maturation of Antigen-presenting cells (APCs), by downmodulating regulatory T cells (Tregs) and decreasing Myeloid-derived suppressor cells (MDSCs) which compose the so-called ‘immune permissive environment’ [high tumor cells to cisplatin [Besides targeting tumor cells, platinum compounds can also elicit an immune response against cancer cells through different mechanisms, i.e., by augmenting CD8ronment’ ,46. Moreronment’ ,49,50,51ronment’ ,53. In kisplatin . Alternaisplatin .Other miRNAs were found functionally involved in modulating cisplatin resistance though the exact molecular mechanisms were unclear ,56,57,58Cisplatin is the standard first-line treatment for advanced-stage NSCLC. It can be administered in combination with third-generation anticancer agents such as Gemcitabine, Docetaxel, Paclitaxel, or Vinorelbine, thus also called platinum-based doublet chemotherapy aka P-doublet . As showMutations in EGFR are frequent in NSCLC patients  and cause the constitutive ligand-independent activation of EGFR receptor thus promoting cell growth and survival . EGFR tyOther pathways were implicated in modulating sensitivity to EGFR-TKIs such as the regulation of NF1 which is a GTPase that negatively regulates the Ras/MEK/ERK pathway and is targeted by miR-641 regulation . Yet, miGefinitib activity can be also modulated by the regulation of miR-146b-5p/nuclear factor kB (NF-kB) axis which impacts the NF-kB-related IL-6 and IL-8 production and ultimately enhances gefitinib-induced apoptosis . Other mOther miRNAs were reported to regulate response to first-generation EGFR-TKIs through direct regulation of newly identified target genes including PELI3 by miR-365a-5p ; ATG5 byEGFR mutations that showed low response to EGFR-TKIs.Further studies are urgently needed to better understand the role of miRNAs in the regulation of response to third-generation TKIs, with a particular focus on atypical Radiotherapy is an important component of the multi-modality treatment for metastatic NSCLC, with ~50% of NSCLC patients undergoing radiotherapy either for a curative or palliative intent . RadiothThe role of miRNA in intrinsic/acquired drug resistance in NSCLC has been mainly investigated using in vitro and in vivo experimental models such as tumor cell lines and mice xenografts and ex vivo samples such as blood and tumor tissues, which were used alone or in combination. Here below, we discuss the most frequently used experimental strategies, highlighting the pros and cons of such methodologies.50) which is the concentration of a drug that induces cell death of 50% of the bulk population. As an alternative, drug response metrics can be expressed as GR50 (drug-induced Growth Rate inhibition) which is less sensitive to growth rate differences influenced, for example, by the cell culture conditions or duration of an experiment [Tumors can be intrinsically resistant to therapy or, alternatively, can become resistant during the course of the treatment (acquired resistance). Overall, we found that a large fraction of studies were performed using in vitro model of acquired resistance which, in our opinion, could represent a potential bias as we later discuss. To get an in vitro model of acquired resistance, commercially available NSCLC cell lines are continuously exposed to increasing high doses of a drug to augment their resistance . Drug reperiment . We founperiment . Yet, thperiment . In addiAs an alternative to acquired resistance models, few other studies performed correlation analysis between drug sensitivity metrics of treatment-naive cells and miRNA expression profile to investigate miRNAs associated with intrinsic drug resistance. In this context, the use of primary cell cultures isolated directly from patient specimens  or alternatively recombinant expression vectors carrying precursor or mature miRNA sequences are both very common strategies to overexpress miRNAs . HoweverContrariwise, miRNA inhibition can be used to knock down a specific miRNA by using for example: (i) antagomirs that are synthetic oligonucleotides designed to be complementary to specific miRNAs, thus preventing their binding to mRNA targets ; (ii) miBesides the identification of a functional role of miRNAs in drug response, some studies have also attempted to identify the exact molecular mechanisms. In Notably, some researchers performed also rescue experiments in which resistance was evaluated upon simultaneous miRNA inhibition and silencing of the target gene or, alternatively, miRNA overexpression and re-expression of the target genes . RecentlIn vivo validation of the miRNA function in the context of the NSCLC therapy is generally performed by taking advantage of immunodeficient mice . Cells are either stably or transiently transfected in vitro with miRNA inhibitors or mimics and injected in mice which are then treated with drugs (xenograft mouse model). Differences in primary tumor growth were then evaluated by using a caliper or, in the case of luciferase reporter constructs, by measuring bioluminescence signals. As an alternative, cell lines can be injected in nude mice which are then treated with the drug alone or in combination with miRNA mimics/inhibitors administered in vivo by intravenous, intraperitoneal, or intratumoral injection. However, a pitfall in this second approach is the impossibility to distinguish whether the biological effect of miRNA expression modulation is exerted exclusively in tumor cells or also in other cell types composing the tumor-microenvironment (TME).Although xenograft models better mimic the tumor microenvironment than models used in in vitro studies, potential limitations arise from the use of cell-line-based xenografts (CDX) mouse models, such as (i) cell lines are continuously propagated in vitro, therefore, after several passages, they may lose resemblance with the original tumor from which they derived; (ii) xenografted cells usually form highly proliferating tumor lesions lacking the architecture of the original tumor and often characterized by hypoxic and/or necrotic regions due to variable perfusions . Thus, tThe lack of a competent immune system in immunodeficient mice is a major limitation when studying the efficacy and/or mechanisms of resistance to immunotherapy . To solvThe role of miRNA in determining drug-resistant phenotypes can be further validated using ex vivo models derived from patient biopsies. The remarkable stability of miRNAs even in harsh conditions such as formalin fixed paraffin-embedded (FFPE) tissue or body fluids  definitely contributed to the success of miRNA expression profile analysis in tumor biopsies ,146. As However, some limitations still apply for example in the bulk tumor miRNA expression profile analysis, which does not provide any clues about miRNA expression profile in specific cell types but rather gives an average expression quantification between tumor cells and TME. Notably, intratumoral biological and genetic heterogeneity bolsters the acquisition of drug resistance because of the presence of a treatment-resistant subpopulation of cells that can outgrow during the course of the treatment. Alternatively, some cells have the ability to survive initial treatment  and acquire de-novo genetic and/or epigenetics alterations that give therapeutic resistance . Such heLastly, ex vivo models may suffer from an additional potential limitation which relies on the use of samples collected mainly from primary lesions instead of metastases. Of course, it is more common during surgery to collect biospecimen from primary lesions than from metastases; indeed, surgery is not an option for advanced metastatic disease . HoweverHere, we made an update on recent research about the functional role of miRNAs in the regulation of resistance to drugs that are currently approved in NSCLC. Overall, miRNAs were more frequently downregulated in drug-resistant than in sensitive samples, and virtually all of the known molecular mechanisms that drive drug resistance in NSCLC cells were found targeted by miRNAs.The number of therapeutic agents approved by the FDA for NSCLC treatment has continuously increased in the last years and, for some drugs, we scored only few or no miRNAs reported to have a functional role. ICIs  have recently become the standard of care for patients with advanced NSCLC disease  thus opeIn conclusion, we note that current limitations in the research field of miRNAs and drug resistance may include: (i) weaknesses of “classical” experimental models such as commercially available 2D in vitro cultures and xenograft mouse models; (ii) intrinsic limitation of methodologies to study miRNA function; (iii) tumor cell heterogeneity and interaction with TME which make the identification of cell-type specific mechanisms involved in drug tolerance complicated. More advanced technologies and experimental models to fully recapitulate the tumor architecture, as well as the molecular and biological heterogeneity of lung cancer, are therefore recommended.A limit of our review could be related to a lack of investigation about the specificity of the miRNAs discussed in terms of eventual similar functions/mechanisms of action in other types of cancers.


The ability to manufacture human hematopoietic stem cells (HSCs) in the laboratory holds enormous promise for cellular therapy of human blood diseases. Several differentiation protocols have been developed to facilitate the emergence of HSCs from human pluripotent stem cells (PSCs). Most approaches employ a stepwise addition of cytokines and morphogens to recapitulate the natural developmental process. However, these protocols globally lack clinical relevance and uniformly induce PSCs to produce hematopoietic progenitors with embryonic features and limited engraftment and differentiation capabilities. This review examines how key intrinsic cues and extrinsic environmental inputs have been integrated within human PSC differentiation protocols to enhance the emergence of definitive hematopoiesis and how advances in genomics set the stage for imminent breakthroughs in this field. Hematopoietic stem cells (HSCs) are defined by their remarkable ability to reconstitute and maintain a functional hematopoietic system over extended periods of time following transplantation into conditioned hosts. This endowment of HSCs to sustain long-term hematopoiesis relates to their ability to either differentiate to produce mature progeny of all myeloid and lymphoid blood cell lineages or self-renew to replace cells that become progressively committed to differentiation. HSCs are thus important cellular targets for the permanent correction of inherited hematologic, metabolic and immunologic disorders. However, allogeneic HSC transplant options are often impeded due to insufficient numbers of transplantable HSCs in donor umbilical cord blood (UCB) units or mobilized peripheral blood (MPB) cell collections, as well as the limited availability of human leukocyte antigen (HLA)-matched donors. Sufficient autologous HSCs for genetic modification are also commonly unavailable in patients with impaired hematopoiesis, such as marrow failure syndromes and chronic inflammatory disorders . Hence, Two general tactics are in development to enhance available sources of HSCs for therapy . Ex vivoAn indirect reprogramming approach that relies on a pluripotent state to change cellular fate was conceptualized in 2006 with Yamanaka’s landmark induction of pluripotency in murine fibroblasts by a defined set of transcription factors  5,6,7].,6,7.5,6,Several basic protocols have been developed to promote hematopoietic differentiation from human pluripotent stem cells. In one approach, iPSCs or ESCs are cultured directly on supportive stromal layers see . ProlongThe terms direct reprogramming, transdifferentiation or conversion have been coined to describe the process of induction of a desired cell lineage from adult somatic cells without transitioning through an intermediary iPSC state . The firWhile direct approaches bypass the complexity of iPSC generation and could, in theory, enable reprogramming of cells in vivo, the efficiency of conversion remains low, safe delivery methods for in vivo transdifferentiation are nonexistent, the limited expansion potential of the post-mitotic starting cell populations may preclude large scale production ex vivo, and the in vitro differentiated cells largely retain an immature phenotype. The indirect approach presents similar pitfalls with modest efficiency of reprogramming and immaturity of the differentiated cellular populations. However, iPSCs can be expanded indefinitely in culture for large-scale clinical production and are readily amenable to genetic manipulation utilizing integrating viral vectors or programmable engineered nuclease technologies. Moreover, the development of a universal, immune-compatible, off-the-shelf product could reduce the inherent costs and slow turnaround time associated with individualized iPSC production, and thus maximize the likelihood of translation to a clinical setting. This review provides a synopsis of recent advances in the indirect reprogramming of human adult somatic cells toward a mesodermal hematopoietic lineage, with an emphasis on therapeutic applications for inherited disorders of the blood system.Transplantability is the conventional criterium to confirm HSC identity after PSC differentiation. To measure human HSC activity, long-term, multilineage engraftment after primary and secondary transplantation into xenograft immune-deficient mouse models represents the current gold standard and is the most stringent assay available. However, the costly and time-consuming in vivo transplantation method is cumbersome and precludes rapid high-throughput assessment of functional HSCs in PSC differentiation cultures. Lymphoid differentiation potential of PSC-derived cells is commonly viewed as an exclusive attribute of HSCs and used as a surrogate for HSC generation ex vivo. However, lympho-myeloid biased progenitors without engraftment potential are known to arise before HSCs in vivo ,28,29,30Approaches to derive HSCs from induced or embryonic pluripotent stem cells in vitro generally attempt to recapitulate the developmental process that occurs during embryonic and fetal development . AdvanceThe process of blood formation during embryonic mammalian development begins with a series of commitment steps within the inner cell mass of the blastocyst. The hypoblast and epiblast layers that form at this stage will later give rise to the yolk sac (YS) and the embryo proper, respectively. A morphological groove termed the primitive streak then appears on the posterior epiblast, marking the commencement of gastrulation ,41,42,43The process of gastrulation results in the formation of a trilaminar germ disc, consisting of the ectoderm, mesoderm and endoderm primordial germ layers. In vivo, the circulatory system  derives from mesodermal cells found at the periphery of the embryo within the splanchnic lateral plate mesoderm. As such, the process of mesodermal induction marks the onset of hematopoiesis in the embryo. Various developmental biology approaches have identified key regulatory networks implicated in this process. Most notably, BMP, FGF, Activin/Nodal and canonical WNT intracellular signaling pathways have distinct and overlapping roles in the induction of mesoderm during development ,44 Figu.Soon after gastrulation, as the developing embryo expands in size and complexity, systemic demands for nutrients, oxygen and waste disposal also evolve. To support the growing fetus, mesodermal progenitors specify hematoendothelial fates, establishing a series of temporally and spatially distinct blood systems designated as “hematopoietic waves”. These successive hematopoietic events first occur in the YS and later within the aorta–gonad–mesonephros (AGM) region of the developing conceptus .The first wave of hematopoiesis, designated as “primitive”, originates in the early YS blood islands. It is marked by the rapid production of large, nucleated red blood cells that arise from mesodermal precursors and can be distinguished by the expression of primitive embryonic globin proteins. This wave also produces unipotent megakaryocyte and macrophage progenitors ,47. The The third wave of hematopoiesis is also labeled “definitive”, but specifically occurs in the embryo proper. It marks the first appearance of multilineage engrafting HSCs primarily within the ventral wall of the dorsal aorta (DA) in the AGM region of the embryo ,50,51,52Vascular endothelial growth factor-A (VEGFA) was identified as a critical initiator signal for the progression of mesoderm to hematoendothelial programs . TranscrHemogenic endothelial cells giving rise to long-term engraftable HSCs have not been associated with venous endothelium during development, suggesting that arterial specification of HE is required for definitive HSC emergence in the AGM ,69,70. AThe mechanism by which HE cells gain hematopoietic potential and morphology at the expense of their endothelial identity during YS and AGM definitive hematopoiesis is known as endothelial-to-hematopoietic transition (EHT) ,84,85. WThe transcription factor RUNX1 is the primary regulator of the EHT process ,54,55,88Recent advances have provided additional insights into the molecular mechanisms that govern endothelial-to-hematopoietic transition. Numerous studies have highlighted the importance of sterile inflammatory signals mediated by the cytokines tumor necrosis factor-α (TNF-α) , interfeThe role of hypoxia and metabolic pathways in response to oxygen concentration and nutrient availability  during EHT has also been underscored in various model systems ,110,111.The regulatory determinants of EHT and HSC emergence continue to expand in complexity, incorporating signaling axes triggered by the biomechanical forces of blood flow ,114,115,Following their emergence within the AGM, nascent HSCs undergo maturation through complex interactions with niche constituents of various hematopoietic sites during development. The functional significance of the AGM region in HSC maturation is supported by numerous studies, but the complexity of the molecular landscape remains to be fully deconvolved ,127,128.Nascent HSCs delaminate from the AGM vascular wall, enter the peripheral circulation and colonize the fetal liver at around 6 weeks of gestation in human embryos, where they undergo considerable expansion and continue to mature in response to microenvironmental cues ,37,129. Definitive HSCs subsequently lodge to the spleen  and boneCurrently employed human PSC differentiation approaches have been largely extrapolated from findings in various model organisms. Most protocols utilize serum-free media and the stepwise addition of cytokines and morphogens to recapitulate in vitro the diverse intrinsic cellular signals known to drive hematopoiesis in vivo. This section examines key intrinsic cues modulated in culture for the de novo generation of human HSCs, with a focus on stage-specific cell signaling pathways, transcription factors (TFs) linked to HSC regulation and epigenetic regulators of hematopoiesis  during development. To bypass constraints of feeder cells and achieve more defined ex vivo culture conditions, natural and synthetic matrices are commonly utilized for the maintenance and expansion of human pluripotent stem cells. Crude ECM extracts derived from murine sarcoma or xeno-free human tissues, such as Matrigel and Geltrex, as well as native or recombinant laminin, collagen, vitronectin, tenascin or fibronectin, key protein components of the ECM, provide a dependable matrix support in PSC culture systems ,244.Although most studies rely on Matrigel coating for hematopoietic differentiation of PSCs , other EAnother report  stemmed Hematopoietic stem cell formation during development is closely regulated by a number of soluble signaling molecules secreted systemically or locally by cellular constituents of microenvironmental niches . Recent In recent years, improved genomics technologies have provided more comprehensive insights into spatial and cell-type-specific gene expression, and enabled the identification of novel factors and extrinsic ligand–receptor pairs during hematopoietic development. One study employed laser capture microdissection coupled with RNA sequencing to map ventrally polarized genes and signaling pathways associated with HSC emergence . GenomicA growing list of soluble regulators have been implicated in HSCs and hematopoietic progenitor cell production during development (reviewed in ), such aAn array of biomechanical inputs contribute to HSC development in the embryo How biomechanical forces can be leveraged for de novo generation of human HSCs will require further investigation. One recent proof-of-concept study used human iPSC-derived HE cells to engineer a novel microfluidic organ-on-a-chip model of the human dorsal aorta and address the impact of flow-related forces on HSC and hematopoietic progenitor cell formation ex vivo . EnhanceThe ability to produce human long-term engrafting HSCs in a dish has far-reaching implications in clinical medicine. Studies in animal and cellular model systems have provided a foundational understanding of hematopoietic ontogeny and, in the past decade, the genomics revolution has delivered new transformative insights that set the stage for imminent breakthroughs in this field. In particular, the recent creation of a single-cell transcriptomic map of human HSC ontogeny now enables the direct comparison of transcriptional similarity between PSC-derived HSCs and distinct native cell types at various stages of hematopoietic maturation. A close transcriptional match between HSCs engineered ex vivo and native immature AGM or extraembryonic HSCs has confirmed their protracted progression to more mature liver and marrow stages in current ex vivo PSC differentiation protocols . Thus, fInadequate activation of core transcription factor networks defining HSC identity could explain, in part, the incomplete maturation and lack of engraftment potential in HSCs generated ex vivo. In previous studies, HSC-specifying TFs utilized to drive PSC differentiation were selected based on evolutionary conservation or enrichment in HSCs compared to more mature progenitors in published gene expression datasets. Recent investigations suggest that genes controlling unique cellular identities are driven by so-called “super-enhancers (SEs)” ,264. ImpEfficient homing, retention and survival within specialized niches of the recipient marrow immediately after transplantation constitute a sine qua non for long-term engraftment of HSCs produced in vitro. An early transplantation failure of PSC-derived HSCs and hematopoietic progenitor cells was uncovered after direct intra-femoral injection in xenograft murine models . The inaMuch work is still required to confirm the potential of these new avenues to comprehensively integrate intrinsic cell signaling and extracellular cues to produce clinical-grade adult-like HSCs. The prospect of this approach to provide durable benefits to human health, however, justifies continued optimism and increasing efforts toward novel scientific advances and optimized ex vivo differentiation schemata of human pluripotent stem cells.


In 2015, the United Nations set important targets to reduce premature cardiovascular disease (CVD) deaths by 33% by 2030. Africa disproportionately bears the brunt of CVD burden and has one of the highest risks of dying from non-communicable diseases (NCDs) worldwide. There is currently an epidemiological transition on the continent, where NCDs is projected to outpace communicable diseases within the current decade. Unchecked increases in CVD risk factors have contributed to the growing burden of three major CVDs—hypertension, cardiomyopathies, and atherosclerotic diseases- leading to devastating rates of stroke and heart failure. The highest age standardized disability-adjusted life years (DALYs) due to hypertensive heart disease (HHD) were recorded in Africa. The contributory causes of heart failure are changing—whilst HHD and cardiomyopathies still dominate, ischemic heart disease is rapidly becoming a significant contributor, whilst rheumatic heart disease (RHD) has shown a gradual decline. In a continent where health systems are traditionally geared toward addressing communicable diseases, several gaps exist to adequately meet the growing demand imposed by CVDs. Among these, high-quality research to inform interventions, underfunded health systems with high out-of-pocket costs, limited accessibility and affordability of essential medicines, CVD preventive services, and skill shortages. Overall, the African continent progress toward a third reduction in premature mortality come 2030 is lagging behind. More can be done in the arena of effective policy implementation for risk factor reduction and CVD prevention, increasing health financing and focusing on strengthening primary health care services for prevention and treatment of CVDs, whilst ensuring availability and affordability of quality medicines. Further, investing in systematic country data collection and research outputs will improve the accuracy of the burden of disease data and inform policy adoption on interventions. This review summarizes the current CVD burden, important gaps in cardiovascular medicine in Africa, and further highlights priority areas where efforts could be intensified in the next decade with potential to improve the current rate of progress toward achieving a 33% reduction in CVD mortality. Approximately three decades ago, conditions such as hypertension and atherosclerotic heart diseases were rare in Africa, and communicable diseases were the major causes of death . HistoriGiven its’ CVD burden, Africa faces several challenges including scarcity of high-quality data, financial constraints, competing priorities, limited skill sets, as well as diagnostic and management challenges . Recent CVDs are the leading cause of morbidity and mortality worldwide, contributing to about a third of all deaths . GloballDrivers of cardiovascular mortality and morbidity in SSA are considerably different compared to HICs. Although ischemic heart disease (IHD) is identified as the leading cause of cardiovascular mortality globally  . NeverthPopulation-based studies in Africa demonstrate a high prevalence of cardiometabolic risk factors with 10-year absolute risk scores ranging 12.5–15.3% for men . HyperteMore than half of CVD deaths in Africa are categorized as premature mortalities, occurring between the ages of 30 and 70 years , 21, 22.The surge in CVD in SSA over the past decade is attributed to a rise in risk factors , largelyThe prevalence of CVD risk factors in Africa has been reported in several heterogeneous studies. The following paragraphs focus on selected classical risk factors with high potential to impact CVD burden in Africa in the foreseeable future .Hypertension (HTN) is described as one of the leading health challenges on the African continent, with an estimated average prevalence of 27% . The repTwenty-four million people are estimated to be living with diabetes on the African continent , a 126% Obesity has been described as a “ticking time bomb” on the African continent, where rates continue to increase rapidly . ObesityAlthough the African region reported the lowest prevalence rates for tobacco smoking (estimated at 18.5%), the rate of decline is low relative to other regions . SeveralHeart failure (HF) is a complex clinical syndrome resulting from a structural or functional impairment of ventricular function . It is tThere are several factors that make HF particularly important in the African context. Foremost, acute HF is the leading cause of admission in specialized cardiology centers , 45. SecIn a systemic review of heart failure in SSA, the contribution of IHD to heart failure etiologies on the continent was small, at 7.2% . HistoriHF management in Africa is still challenged by lack of basic supportive investigations such as electrocardiograms, chest x-ray, and echocardiography, which are available in less than 50% of diagnosing centers , 53. TheThe high rate of preventable heart failure should draw attention to prevention as an important area of focus for the continent. Other efforts should include investing in high quality epidemiological research -including therapeutic interventions, prioritizing early diagnosis, and optimizing treatments according to evidence-based medicine. Further, innovative strategies such as tech-enabled ways to enhance patient follow-up and promote adherence may lead to improved outcomes.Sub-Saharan Africa has witnessed a substantial increase of atherosclerotic cardiovascular diseases (ASCVD) over the last three decades. According to the Global Burden of Disease (GBD) estimates, the highest contribution to CVD burden on the continent is attributed to atherosclerotic diseases, with 71.4, 37.7, and 154% increases in the burden  of ischemic heart disease, stroke and peripheral artery disease since 1990 . While tCoronary artery disease (CAD) is the leading cause of death globally, responsible for over 50% of all cardiovascular deaths . Over thAfrican countries face several challenges with regards to CAD. Apart from poor recognition of angina symptoms , there iAs with CAD, there has been a substantial decline in global, age-standardized stroke mortality rates over the last two decades, possibly as a result of improved stroke care. However, this decline in incidence has not been reflected in Africa . The annInadequate control of stroke risk factors, poor recognition of the urgency of symptoms leading to delayed presentation, lack of timely diagnosis and fibrinolytic services, and the dearth of post-stroke rehabilitation services are major drivers of stroke burden in SSA , 68. It Moving forward, investment in cost-effective ways for the prevention and treatment of risk factors for atherosclerotic disease is paramount. Systematic screening, improving access to medication, and implementing standardized guideline-directed clinical management may prove vital to control efforts . InvestiHypertension is the number one driver of CVD in Africa. According to recent estimates, the African region has the highest prevalence of hypertension in the world , as wellDubbed the “silent killer,” a result of the initial asymptomatic nature of the disease, many present for the first time to health care facilities with fatal complications. Hypertension is responsible for more than half of total cardiovascular related deaths, being the main driver of IHD, HHD, and stroke burden on the continent . In a 3-The benefits of adequately controlled blood pressure are well-known, with reported reductions in cardiovascular mortality as large as 38.0 and 30.4% among females and males . Of critThe need for hypertension control in Africa is undoubtedly one of the highest priorities. Integrating care widely in primary health facilities by equipping them with tools and quality, accessible medications to manage hypertension in concert with other cardiovascular risk factors should be a priority . One appSub-Saharan Africa (SSA) has the highest burden of HIV in the world, accounting for more than two thirds of the global HIV burden . RemarkaIn high-income countries, people living with HIV (PLHIV) and hypertension have higher risks of cardiovascular events and all-cause mortality compared to HIV-uninfected persons with hypertension or PLHIV with normal blood pressure . AbsolutIntegrating CVD care into HIV programs is key to mitigating the growing burden of CVD in HIV populations in Africa. As an example, PLHIV attending primary health care facilities in rural South Africa received better care for both hypertension and diabetes . But in Cardiomyopathies are defined as disorders in which the heart muscle is structurally and functionally abnormal, in the absence of CAD, hypertension, valvular disease, and congenital heart disease (CHD) sufficient to cause the observed myocardial abnormality . They arDCM is defined by the presence of left ventricular dilatation and left ventricular systolic dysfunction in the absence of abnormal loading conditions  or coronary artery disease sufficient to cause global systolic impairment . DCM is Two thirds of patients with DCM have poor prognostic factors and die within 5 years of their first symptoms . The cauA number of other cardiomyopathies contribute to CVDs in Africa, albeit much less common than DCM. Hypertrophic cardiomyopathy (HCM) was hardly diagnosed in Africa prior to widespread availability of echocardiography . CurrentEndomyocardial fibrosis (EMF), an endemic cardiomyopathy, is believed to be the most common primary form of restrictive cardiomyopathy in Africa, elaborated on later in this review. Other rare forms of cardiomyopathies which have been reported in Africa include ARVC and NCLV (non-compaction of the left ventricle). The diagnosis of ARVC is still a challenge in SSA countries. The only available data on this condition comes from a single center study in South Africa, where 12 cases were reported in 6 years . NCLV isOverall, investments into further research will be informative in enhancing our understanding of the etiology and management of cardiomyopathies in Africa. Lack of confirmatory diagnostic facilities, i.e., endomyocardial biopsy, cardiac magnetic resonance imaging, and genetic studies have contributed to under-diagnosis of some cardiomyopathies in Africa; however, the rising use of echocardiography has increased suspicion and may result in increased reporting in the coming years.RHD results from recurrent episodes of acute rheumatic fever (ARF), a long-term complication of Group A streptococcal (GAS) infection. ARF is thought to be an autoimmune-mediated response to interactions between the body and certain components of the GAS bacterial cell wall protein. RHD is currently estimated to affect approximately forty million people worldwide, the majority of whom are children and young adults in sub-Saharan Africa . About 3A combination of factors including health system factors, poor health seeking behavior, and evolving virulence of the causative agent have resulted in the persistence of GAS and its long term sequelae, RHD, in SSA . The burPrimordial prevention involves a multi-pronged approach at the government level that targets improvements in living conditions of the population including improving housing standards to prevent overcrowding, fostering good hygiene practices, and promoting education. Primary prevention aims at preventing the rheumatic process from starting and targets prompt detection, treatment, and follow up of GAS pharyngitis. Although several interventions have been implemented in Cuba and Nicaragua, this approach requires near overhauling of the health system and may not be cost effective (Secondary prevention that targets reducing recurrent episodes of ARF and thus RHD progression is by far the most effective intervention to date. A registry-based approach based on monthly injectable benzathine penicillin (BPG) has been shown to be both effective and affordable  How can health systems in LMIC improve access to primary prevention services? (2) In patients with ARF or RHD, which delivery systems can help improve adherence to BPG medication? Are depot, long term preparations of penicillin feasible? (3) Does ARF present differently in Africa and are health care workers adequately equipped to diagnose, manage and perform its surveillance? And (4) What are the immunological processes that underlie progression from ARF to clinical RHD? Several important research works are on-going examining some of the aforementioned aspects of RHD management. The landmark GOAL trial showed that BPG prevented progression of latent RHD in children who were administered monthly BPG . FurtherThe CVD epidemic in Africa includes and/or is a harbinger for cardiac arrhythmias and their related health impact . The speChief among the cardiac arrhythmia maladies is atrial fibrillation (AF), noted to have a higher prevalence in the general population than previously thought . A recenLikewise, data on the epidemiology and management of supraventricular arrhythmias, ventricular arrhythmias, brady-arrhythmias and sudden cardiac arrest in Africa remain scant . The AfrOverall, there is a growing need for appropriate arrhythmia care in Africa, an area that remains largely unmet because of complex multi-level challenges . LocallyEndomyocardial fibrosis (EMF) is the most common form of restrictive cardiomyopathy globally . Since iThe etiopathogenetic mechanisms underlying the disease still remain debatable, in part due to lack of investment in research. Environmental , socioeconomic, and genetic factors are all thought to play a role , 125–127Advanced EMF carries a poor prognosis. Medical therapy is directed toward treatment of heart failure symptoms. Death often results from refractory heart failure, arrhythmias and thromboembolism. Corrective surgery, which has been shown to improve survival and quality of life , 129, isIn patients with diagnosed and undiagnosed CVD, pregnancy and labor pose remarkable physiological and hemodynamic stress and portends significant negative consequences on both maternal and fetal outcomes. Globally, 1–4% of pregnancies are complicated by maternal CVDs (excluding hypertension) . On its A systematic review on antenatal heart disease burden in South Africa showed a prevalence of 0.6% . WhereasChallenges still exist in the management of patients with heart disease in pregnancy. African women in general are under social pressure to bear children and will take the risk despite the severity of disease . SecondlImproving outcomes of pregnant mothers with CVD requires an integrated and context- specific approach that engages the patient (understanding the patient needs), and involves a multi-disciplinary team headed by an obstetrician and cardiologist. Joint obstetric-cardiac care has been shown to improve both maternal and fetal outcomes . SustainCongenital heart disease accounts for one third of all birth defects . The gloSeveral challenges affect CHD care in Africa: (1) prenatal diagnosis of CHD in Africa is rare because of severely limited antenatal screening capacity for CHD , 157. 2. (3) AccTo improve CHD care, it is important to: (1) Increase funding to strengthen already existing CHD centers through both governmental and non-governmental organizations, (2) support training and mentorship of super-specialists across the CHD spectrum, and (3) increase awareness, promote early neonatal screening, and treatment through universal health insurance.status quo projections indicate that Africa’s overall progress is insufficient to achieve SDG 3.4 by 2030 (Currently half way between 2015 and 2030,  by 2030 . Data fr by 2030 . Figure  by 2030 . These p•There’s inadequate research productivity from Africa which contributes to the widening gap in health outcomes with other countries , 168. Im•Data on the physician-to-population ratio confirm a persistent human resource crisis in the health sector, which is even more pronounced for sub-specialties . Overtim•There is a large gap to fill for general access to CVD care, contributory factors include:∘Low health budget allocation—According to the WHO, most African countries have insufficient health financing systems with high out of pocket expenditures. Almost half of all African countries still have out of pocket expenditures of above 40% . In gene∘Lack of universal health insurance and consequently high out-of-pocket expenditure, sometimes resulting in catastrophic health spending.∘Inadequate access and long-term affordability of evidenced-based CVD medications, including important drugs for CVD prevention .∘Inadequate facilities for advanced cardiovascular imaging and procedures .•Low education and health literacy levels , affects•Prevention: identifying and implementing cost effective, preventive strategies targeting both high risk and the general population is the most reliable solution to the rising burden of CVD in Africa. Preventive efforts such as the adoption of the FCTC for tobacco control ( control , introdu control , and impstatus quo (Hypertension Prevention deserves concerted efforts given its contribution to CVD in Africa. Universal and periodic screening, diagnosis and guideline-directed treatment through strengthening and integration into primary care facilities, task-shifting, and universal availability of effective drugs should be prioritized in most countries. Intensively accelerating control through two population-based interventions—pharmacological guideline-directed treatment of hypertension and “upstream” policies targeted at a reduction in salt consumption could reduce CVD mortality by up to 22% relative to atus quo . The WHOatus quo .•Increase Research Outputs: high-quality epidemiological and implementation research, as well as dedicated investments toward strengthening vital statistic data and systematic surveillance, will inform practice and contribute towards identifying and evaluating’ sustainable interventions in various settings (settings , track p•Improve Training: priorities to improve training should include establishing in-country cardiology training centers and strengthening existing training centers through bench marking and fostering collaboration with strong external cardiology training institutions.•Improve Access: a number of options exists to increase access to cardiovascular care:∘reducing out-of-pocket expenditure and implementing health financing plans with the objective of increasing equitable and effective access to CVD care . Among o∘task shifting of preventive cardiovascular care strategies like screening, lifestyle modification counseling, routine, uncomplicated, follow-ups, specialized care linkage, and coordination of NCD support groups .∘investing in infrastructure for high-quality care including acute cardiovascular care is an essential component of reducing cardiovascular mortality .∘incentives for health care workers that aim to enhance service delivery and retain knowledgeable and skilled medical personnel within countries .The burden of CVD in Africa is a result of increasing and unchecked risk factors with considerable heterogeneity in risk trends across its regions, that has led to explosive rates of CVD-related morbidity and mortality on the continent. Despite the sustained efforts to combat CVDs on the continent by the different advocacy groups such as WHO, PASCAR, and World Heart Federation, the adoption of effective policies is lagging in many African states. This review has highlighted several gaps in various sectors of CVD care on the continent. Overall, the evidence suggests applying preventive strategies to whole populations that target major risk factors will make important contributions toward reaching set targets by 2030. Specifically, for Africa, intensified and tailored regional efforts need to be channeled to lower blood pressure and other risk factors that should be coupled with continued surveillance and reliable data collection and monitoring programs. Concerted efforts from global, regional, and local experts are needed to increase advocacy for CVDs. Efforts should focus on identifying innovative ways to improve access and service provision at a primary health care level, investing toward universal health coverage, investing in cost-effective preventive approaches that are easily adaptable, and prioritizing research. Given country commitments to strengthen health systems, adopt and implement key metrics targeting awareness, prevention and management of CVDs, significant strides are possible toward changing the current trajectory of CVD burden in Africa.EO, JR, DN, and NM contributed to conception and initial outline of the review. NM, DN, TA, WZ, IS, JN, WA, SL, EN, JR, EO, and JK contributed to writing different subsections. JK, EO, EN, SL, DN, and NM contributed to manuscript content and language editing. All authors contributed to manuscript revision, read, and approved the submitted version.


Alzheimer’s disease and related dementias are prevalent, incurable, and highly impactful diagnoses. Dementias are therefore feared diagnoses. Dementia-related anxiety (DRA) is anxiety about a current or future diagnosis of dementia and the associated complex symptoms. In a mixed methods study, semi-structured interviews were conducted to identify causes of DRA and revealed that numerous adults anticipated suicidal or death ideation if diagnosed with dementia. Fifty cognitively healthy, community-dwelling adults aged 58 to 89  were recruited from a university participant registry and Memory Clinic. Among participants endorsing anticipated suicidal or death ideation, responses ranged from active plans, including interest in physician-assisted suicide, to more passive wishes to hasten death rather than continue to live with dementia. Within reports of both anticipated suicidal and death ideation, subthemes emerged, including the concern about becoming a burden to others in more advanced stages of dementia, the devaluation of life or the self with dementia, and the desire for (and anticipated thwarting of) control and independence. Statements of anticipated suicidal and death ideation were contingent on a future dementia diagnosis and may reflect errors in affective forecasting. Nevertheless, given the prevalence of dementias and older adults’ elevated rates of suicide, the intersection of these two public health issues warrants greater attention and further investigation.