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Platelets are the terminal progeny of megakaryocytes, primarily produced in the bone marrow, and play critical roles in blood homeostasis, clotting, and wound healing. Traditionally, megakaryocytes and platelets are thought to arise from multipotent hematopoietic stem cells (HSCs) via multiple discrete progenitor populations with successive, lineage-restricting differentiation steps. However, this view has recently been challenged by studies suggesting that (1) some HSC clones are biased and/or restricted to the platelet lineage, (2) not all platelet generation follows the "canonical" megakaryocytic differentiation path of hematopoiesis, and (3) platelet output is the default program of steady-state hematopoiesis. Here, we specifically investigate the evidence that in vivo lineage tracing studies provide for the route(s) of platelet generation and investigate the involvement of various intermediate progenitor cell populations. We further identify the challenges that need to be overcome that are required to determine the presence, role, and kinetics of these possible alternate pathways.

Epigenetic mechanisms regulate the multilineage differentiation capacity of hematopoietic stem cells (HSCs) into a variety of blood and immune cells. Mapping the chromatin dynamics of functionally defined cell populations will shed mechanistic insight into 2 major, unanswered questions in stem cell biology: how does epigenetic identity contribute to a cell type's lineage potential, and how do cascades of chromatin remodeling dictate ensuing fate decisions? Our recent work revealed evidence of multilineage gene priming in HSCs, where open cis-regulatory elements (CREs) exclusively shared between HSCs and unipotent lineage cells were enriched for DNA binding motifs of known lineage-specific transcription factors. Oligopotent progenitor populations operating between the HSCs and unipotent cells play essential roles in effecting hematopoietic homeostasis. To test the hypothesis that selective HSC-primed lineage-specific CREs remain accessible throughout differentiation, we used ATAC-seq to map the temporal dynamics of chromatin remodeling during progenitor differentiation. We observed epigenetic-driven clustering of oligopotent and unipotent progenitors into distinct erythromyeloid and lymphoid branches, with multipotent HSCs and MPPs associating with the erythromyeloid lineage. We mapped the dynamics of lineage-primed CREs throughout hematopoiesis and identified both unique and shared CREs as potential lineage reinforcement mechanisms at fate branch points. Additionally, quantification of genome-wide peak count and size revealed overall greater chromatin accessibility in HSCs, allowing us to identify HSC-unique peaks as putative regulators of self-renewal and multilineage potential. Finally, CRISPRi-mediated targeting of ATACseq-identified putative CREs in HSCs allowed us to demonstrate the functional role of selective CREs in lineage-specific gene expression. These findings provide insight into the regulation of stem cell multipotency and lineage commitment throughout hematopoiesis and serve as a resource to test functional drivers of hematopoietic lineage fate.

Introduction: Collaborative robots, designed to work alongside humans for manipulating end-effectors, greatly benefit from the implementation of active constraints. This process comprises the definition of a boundary, followed by the enforcement of some control algorithm when the robot tooltip interacts with the generated boundary. Contact with the constraint boundary is communicated to the human operator through various potential forms of feedback. In fields like surgical robotics, where patient safety is paramount, implementing active constraints can prevent the robot from interacting with portions of the patient anatomy that shouldn’t be operated on. Despite improvements in orthopaedic surgical robots, however, there exists a gap between bulky systems with haptic feedback capabilities and miniaturised systems that only allow for boundary control, where interaction with the active constraint boundary interrupts robot functions. Generally, active constraint generation relies on optical tracking systems and preoperative imaging techniques.Methods: This paper presents a refined version of the Signature Robot, a three degrees-of-freedom, hands-on collaborative system for orthopaedic surgery. Additionally, it presents a method for generating and enforcing active constraints “on-the-fly” using our previously introduced monocular, RGB, camera-based network, SimPS-Net. The network was deployed in real-time for the purpose of boundary definition. This boundary was subsequently used for constraint enforcement testing. The robot was utilised to test two different active constraints: a safe region and a restricted region.Results: The network success rate, defined as the ratio of correct over total object localisation results, was calculated to be 54.7% ± 5.2%. In the safe region case, haptic feedback resisted tooltip manipulation beyond the active constraint boundary, with a mean distance from the boundary of 2.70 mm ± 0.37 mm and a mean exit duration of 0.76 s ± 0.11 s. For the restricted-zone constraint, the operator was successfully prevented from penetrating the boundary in 100% of attempts.Discussion: This paper showcases the viability of the proposed robotic platform and presents promising results of a versatile constraint generation and enforcement pipeline.

Hematopoietic stem cell (HSC) multipotency and self-renewal are typically defined through serial transplantation experiments. Host conditioning is necessary for robust HSC engraftment, likely by reducing immune-mediated rejection and by clearing limited HSC niche space. Because irradiation of the recipient mouse is non-specific and broadly damaging, there is a need to develop alternative models to study HSC performance at steady-state and in the absence of radiation-induced stress. We have generated and characterized two new mouse models where either all hematopoietic cells or only HSCs can be specifically induced to die in vivo or in vitro. Hematopoietic-specific Vav1-mediated expression of a loxP-flanked diphtheria-toxin receptor (DTR) renders all hematopoietic cells sensitive to diphtheria toxin (DT) in "Vav-DTR" mice. Crossing these mice to Flk2-Cre mice results in "HSC-DTR" mice which exhibit HSC-selective DT sensitivity. We demonstrate robust, rapid, and highly selective cell ablation in these models. These new mouse models provide a platform to test whether HSCs are required for long-term hematopoiesis in vivo, for understanding the mechanisms regulating HSC engraftment, and interrogating in vivo hematopoietic differentiation pathways and mechanisms regulating hematopoietic homeostasis.

Although WNT signaling is frequently dysregulated in solid tumors, drugging this pathway has been challenging due to off-tumor effects. Current clinical pan-WNT inhibitors are nonspecific and lead to adverse effects, highlighting the urgent need for more specific WNT pathway-targeting strategies. We identified elevated expression of the WNT receptor Frizzled class receptor 7 (FZD7) in multiple solid cancers in The Cancer Genome Atlas, particularly in the mesenchymal and proliferative subtypes of ovarian serous cystadenocarcinoma, which correlate with poorer median patient survival. Moreover, we observed increased FZD7 protein expression in ovarian tumors compared with normal ovarian tissue, indicating that FZD7 may be a tumor-specific antigen. We therefore developed a novel antibody-drug conjugate, septuximab vedotin (F7-ADC), which is composed of a chimeric human-mouse antibody to human FZD7 conjugated to the microtubule-inhibiting drug monomethyl auristatin E (MMAE). F7-ADC selectively binds human FZD7, potently kills ovarian cancer cells in vitro, and induces regression of ovarian tumor xenografts in murine models. To evaluate F7-ADC toxicity in vivo, we generated mice harboring a modified Fzd7 gene where the resulting Fzd7 protein is reactive with the human-targeting F7-ADC. F7-ADC treatment of these mice did not induce acute toxicities, indicating a potentially favorable safety profile in patients. Overall, our data suggest that the antibody-drug conjugate approach may be a powerful strategy to combat FZD7-expressing ovarian cancers in the clinic.

Hematopoietic stem cells (HSCs) have been studied extensively since their initial functional description in 1961 when Dr. James Till and Dr. Ernest McCulloch developed the first in vivo clonal strategy, termed the spleen colony-forming unit (CFU-S) assay, to assess the functional capacity of bone marrow-derived hematopoietic progenitors at the single-cell level. Through transplantation of bone marrow cells and analysis of the resulting cellular nodules in the spleen, the CFU-S assay revealed both the self-renewal and clonal differentiation capacity of hematopoietic progenitors. Further development and use of this assay have identified highly proliferative, self-renewing, and differentiating HSCs that possess clonal, multilineage differentiation. The CFU-S strategy has also been adapted to interrogating single purified hematopoietic stem and progenitor cell populations, advancing our knowledge of the hematopoietic hierarchy. In this review, we explore the major discoveries made with the CFU-S assay, consider its modern use and recent improvements, and compare it with commonly used long-term transplantation assays to determine the continued value of the CFU-S assay for understanding HSC biology and hematopoiesis.

Though WNT signaling is frequently dysregulated in solid tumors, drugging this pathway has been challenging due to off-tumor effects. Current clinical pan-WNT inhibitors are non-specific and lead to adverse effects, highlighting the urgent need for more specific WNT-pathway targeting strategies. We identified elevated expression of the WNT receptor Frizzled class receptor 7 (FZD7) in multiple solid cancers in The Cancer Genome Atlas, particularly in the mesenchymal and proliferative subtypes of ovarian serous cystadenocarcinoma, which correlate with poorer median patient survival. Moreover, we observed increased FZD7 protein expression in ovarian tumors compared to normal ovarian tissue, indicating that FZD7 may be a tumor-specific antigen. We therefore developed a novel antibody-drug conjugate, septuximab vedotin (F7-ADC), which is composed of a chimeric human-mouse antibody to human FZD7 conjugated to the microtubule-inhibiting drug monomethyl auristatin E (MMAE). F7-ADC selectively binds human FZD7, potently kills ovarian cancer cells in vitro, and induces regression of ovarian tumor xenografts in murine models. To evaluate F7-ADC toxicity in vivo, we generated mice harboring a modified Fzd7 gene where the resulting Fzd7 protein is reactive with the human-targeting F7-ADC. F7-ADC treatment of these mice did not induce acute toxicities, indicating a potentially favorable safety profile in patients. Overall, our data suggest that the antibody-drug conjugate approach may be a powerful strategy to combat FZD7-expressing ovarian cancers in the clinic.

Aim: This case report aims to illustrate a clinical protocol that allows for the rehabilitation of patients requiring extensive osteotomy, simultaneous implant placement, and full-arch, screwed-in prosthetics in one session. This protocol allows for the improvement of the aesthetics and functionality of the fixed implant-supported prosthesis through the preoperative planning of all surgical procedures, including osteotomy, and of the prosthesis through the application of 3D-printing technology for the creation of surgical templates and prostheses. Methods: This case report concerns a 72-year-old patient, ASA1, who, following diagnosis, the establishment of a treatment plan, and the provision of informed consent, opted for an immediate, full-arch rehabilitation of the lower arch. The digital planning stage started with the correct positioning of the fixtures. The proper bone levels were found and used to guide the creation of the provisional screwed-in prothesis. Two templates with the same supports (landmarks/pins) were then 3D-printed: a positioning template, including a slit to assist the surgeon during the osteotomy, and a surgery template to assist the surgeon during the implants’ positioning. A screwed-in prosthesis encased in resin C&B MFH (NEXTDENT®, Soesterberg, The Netherlands) was delivered. Minimal occlusal adjustments were performed. Results: In a single clinical session, through careful planning and the pre-operative 3D printing of a prosthesis, a temporary implant-supported prosthetic rehabilitation was possible in a case that required an extended osteotomy. Clinically, the correspondence between the virtual design phase and the final realization was consistent. At a functional level, the provisional prosthesis required minimal occlusal adjustments and the DVO values obtained in the immediate post-operative period were found to be comparable to those of the virtual design. By planning the final position of the bone and the implants in advance, it was possible to deliver a full-arch prothesis with proper implant emergence, occlusal vertical dimensions, and occlusal relationship. Conclusion: This fully digital protocol allows the clinician to preview and plan the osteotomy and implant surgery as well as the delivery of the temporary, immediately loaded, complete, fixed prosthesis in patients who are candidates for post-extraction surgery with the need for severe osteotomy.

Platelets are the terminal progeny of megakaryocytes, primarily produced in the bone marrow, and play critical roles in blood homeostasis, clotting, and wound healing. Traditionally, megakaryocytes and platelets are thought to arise from multipotent hematopoietic stem cells (HSCs) via multiple discrete progenitor populations with successive, lineage-restricting differentiation steps. However, this view has recently been challenged by studies suggesting that (1) some HSC clones are biased and/or restricted to the platelet lineage, (2) not all platelet generation follows the “canonical” megakaryocytic differentiation path of hematopoiesis, and (3) platelet output is the default program of steady-state hematopoiesis. Here, we specifically investigate the evidence that in vivo lineage tracing studies provide for the route(s) of platelet generation and investigate the involvement of various intermediate progenitor cell populations. We further identify the challenges that need to be overcome that are required to determine the presence, role, and kinetics of these possible alternate pathways.

Imaging and reconstruction of sheep brain axonal cytoarchitecture provides insight for brain biomechanics models that simulate drug delivery and other biological processes governed by interstitial fluid flow and transport.

Aim: The conometric concept was proposed as a possible connection between the abutment and the prosthetic coping. This research aimed to review the features and possible clinical uses of this connection in an implant-supported fixed prosthesis. Methods: An electronic search was conducted on an online database for the topic in object; articles published in international literature were considered and the research gave 17 results, and 6 parameters were analyzed. Results: This connection eliminated the possibility of cement residues in the subgingival region, reducing the risk of inflammation of peri-implant soft and hard tissues; not having to remove the cement residues, it is possible to place the margins in more apical portions, improving the aesthetics outcomes of the rehabilitations. It is also known that the retention by means of a screw causes a weakening of the restoration. The retentive force is adequate for fixed rehabilitation even after a high number of insertion–disengagement cycles; in vitro studies have also shown a high bacterial sealing. Implant rehabilitation using preformed components, such as conometric hoods, is helpful for CAD/CAM, so a digital workflow is possible. Several types of prosthesis were presented, all of which demonstrated adequate clinical performance in the follow-up observation. Conclusions: This type of connection seems to be suitable to support fixed implant rehabilitations, but long-term clinical studies are needed to validate this system.

The digital transformation has revolutionized various sectors, including dentistry. Dentistry has emerged as a pioneer in embracing digital technologies, leading to advancements in surgical and prosthetic oral healthcare. Immediate loading for full-arch edentulous dental implants, once debated, is now widely accepted. This case report describes a 74-year-old patient with dental mobility and significant bone loss who was rehabilitated using a Toronto Bridge protocol on four dental implants with immediate loading. Digital planning, surgical guides, 3D printing, and precision techniques were employed. The surgery involved implant placement and prosthetic procedures. The patient reported minimal post-operative discomfort, and after four months, the definitive prosthesis was successfully placed. This case demonstrates the efficacy of immediate loading in complex dental scenarios with digital innovation, resulting in improved patient outcomes. The full digital workflow, including 3D printing and the use of modern materials, enhances the efficiency and predictability of oral rehabilitation, marking a transformative era in dental care. The integration of digital technology in all phases of treatment, from diagnosis to finalization, makes this approach safer, reliable, and efficient, thereby benefiting both patients and clinicians.

Abstract An automatic markerless knee tracking and registration algorithm has been proposed in the literature to avoid the marker insertion required by conventional computer-assisted knee surgery, resulting in a shorter and less invasive surgical workflow. However, such an algorithm considers intact femur geometry only. The bone surface modification is inevitable due to intra-operative intervention. The mismatched correspondences will degrade the reliability of registered target pose. To solve this problem, this work proposed a supervised deep neural network to automatically restore the surface of processed bone. The network was trained on a synthetic dataset that consists of real depth captures of a model leg and simulated realistic femur cutting. According to the evaluation on both synthetic data and real-time captures, the registration quality can be effectively improved by surface reconstruction. The improvement in tracking accuracy is only evident over test data, indicating the need for future enhancement of the dataset and network.

Abstract Hematopoietic stem cell (HSC) multipotency and self-renewal are typically defined through serial transplantation experiments. Host conditioning is necessary for robust HSC engraftment, likely by reducing immune-mediated rejection and by clearing limited HSC niche space. Because irradiation of the recipient mouse is non-specific and broadly damaging, there is a need to develop alternative models to study HSC performance at steady-state and in the absence of radiation-induced stress. We have generated and characterized two new mouse models where either all hematopoietic cells or only HSCs can be specifically induced to die in vivo or in vitro. Hematopoietic-specific Vav1-mediated expression of a loxP-flanked diphtheria-toxin receptor (DTR) renders all hematopoietic cells sensitive to diphtheria toxin (DT) in “Vav-DTR” mice. Crossing these mice to Flk2-Cre mice results in “HSC-DTR” mice which exhibit HSC-selective DT sensitivity. We demonstrate robust, rapid, and highly selective cell ablation in these models. These new mouse models provide a platform to test whether HSCs are required for long-term hematopoiesis in vivo, for understanding the mechanisms regulating HSC engraftment, and interrogating in vivo hematopoietic differentiation pathways and mechanisms regulating hematopoietic homeostasis.

This investigation aimed to compare the effectiveness of the OT Equator® (Rhein, Bologna, Italy) and the Locator attachment systems used to retain early loaded implant-retained overdentures. This study was designed as a multicenter randomised controlled trial of parallel groups. After implant placement, the patients were randomised to receive OT Equator® attachments in the test group or Locator attachments in the control group. The outcome measures were implant and prosthetic success and survival rates, any biological and technical complication, marginal bone loss, patients’ satisfaction, and periodontal parameters. Overall, 42 patients were consecutively enrolled and treated. One implant was lost in the control group, while no implants were lost in the test group. No prostheses failed in both groups. Only a few complications were experienced in both groups. The main was represented by loss of retention of the attachments (retentive caps). The OT Equator® attachment showed statistically lower periodontal parameters. In conclusion, both attachment systems were suitable for overdenture implant retention.