Homo sapiens and Neanderthals share high cerebral cortex integration into adulthood.

There is controversy around the mechanisms that guided the change in brain shape during the evolution of modern humans. It has long been held that different cortical areas evolved independently from each other to develop their unique functional specializations. However, some recent studies suggest that high integration between different cortical areas could facilitate the emergence of equally extreme, highly specialized brain functions. Here, we analyse the evolution of brain shape in primates using three-dimensional geometric morphometrics of endocasts. We aim to determine, firstly, whether modern humans present unique developmental patterns of covariation between brain cortical areas; and secondly, whether hominins experienced unusually high rates of evolution in brain covariation as compared to other primates. On the basis of analyses including modern humans and other extant great apes at different developmental stages, we first demonstrate that, unlike our closest living relatives, Homo sapiens retain high levels of covariation between cortical areas into adulthood. Among the other great apes, high levels of covariation are only found in immature individuals. Secondly, at the macro-evolutionary level, our analysis of 400 endocasts, representing 148 extant primate species and 6 fossil hominins, shows that strong covariation between different areas of the brain in H. sapiens and Homo neanderthalensis evolved under distinctly higher evolutionary rates than in any other primate, suggesting that natural selection favoured a greatly integrated brain in both species. These results hold when extinct species are excluded and allometric effects are accounted for. Our findings demonstrate that high covariation in the brain may have played a critical role in the evolution of unique cognitive capacities and complex behaviours in both modern humans and Neanderthals.

What Do Underrepresented in Medicine Junior Family Medicine Faculty Value From a Faculty Development Experience?

BACKGROUND AND OBJECTIVES: While there is increased attention to underrepresented in medicine (URiM) faculty and students, little is known about what they value in faculty development experiences. METHODS: We performed a URiM-focused, 3-day family medicine faculty development program and then collected program evaluation forms. The program evaluations had open-ended questions and a reflection on the activity. We used inductive open coding using NVivo software. We analyzed open-ended responses and reflections, and identified themes. RESULTS: Seven participants provided reflections on the workshop and responses to the evaluation forms. Analysis revealed four major themes in the learners' responses and reflections: (1) personalizing learning, (2) impacting career trajectories, (3) clarifying the writing process, and (4) creating a safe place, with frequencies of 28.2%, 26.7%, 23.6%, and 20.9%, respectively. CONCLUSIONS: Although this faculty development experience was designed to teach writing skills to URiM junior faculty, their collective responses indicate that they found value beyond the skills taught and appreciated the approach taken in this activity.

Towards a common lexicon for equity, diversity, and inclusion work in academic medicine.

Differential rewarding of work and experience has been a longtime feature of academic medicine, resulting in a series of academic disparities. These disparities have been collectively called a cultural or minority "tax," and, when considered beyond academic medicine, exist across all departments, colleges, and schools of institutions of higher learning-from health sciences to disciplines located on university campuses outside of medicine and health. A shared language can provide opportunities for those who champion this work to pool resources for larger impacts across the institution. This article aims to catalog the terms used across academic medicine disciplines to establish a common language describing the inequities experienced by Black, Latinx, American Indian/Alaska Native and Native Hawaiian/Other Pacific Islander, Women, and other underrepresented people as well as queer, disabled, and other historically marginalized or excluded groups. These ideas are specific to academic medicine in the United States, although many can be used in academic medicine in other countries. The terms were selected by a team of experts in equity, diversity, and inclusion, (EDI) who are considered national thought leaders in EDI and collectively have over 100 years of scholarship and experience in this area.

Relative Brain Volume of Carnivorans Has Evolved in Correlation with Environmental and Dietary Variables Differentially among Clades.

Carnivorans possess relatively large brains compared to most other mammalian clades. Factors like environmental complexity (Cognitive Buffer Hypothesis) and diet quality (Expensive-Tissue Hypothesis) have been proposed as mechanisms for encephalization in other large-brained clades. We examine whether the Cognitive Buffer and Expensive-Tissue Hypotheses account for brain size variation within Carnivora. Under these hypotheses, we predict a positive correlation between brain size and environmental complexity or protein consumption. Relative endocranial volume (phylogenetic generalized least-squares residual from species' mean body mass) and 9 environmental and dietary variables were collected from the literature for 148 species of terrestrial and marine carnivorans. We found that the correlation between relative brain volume and environment and diet differed among clades, a trend consistent with other larger brained vertebrates (i.e., Primates, Aves). Mustelidae and Procyonidae demonstrate larger brains in species with higher-quality diets, consistent with the Expensive-Tissue Hypothesis, while in Herpestidae, correlations between relative brain size and environment are consistent with the Cognitive Buffer Hypothesis. Our results indicate that carnivorans may have evolved relatively larger brains under similar selective pressures as primates despite the considerable differences in life history and behavior between these two clades.

An Approach to Faculty Development for Underrepresented Minorities in Medicine.

OBJECTIVES: The diversity of the US physician workforce lags significantly behind the population, and the disparities in academic medicine are even greater, with underrepresented in medicine (URM) physicians accounting for only 6.8% of all US medical school faculty. We describe a "for URM by URM" pilot approach to faculty development for junior URM Family Medicine physicians that targets unique challenges faced by URM faculty. METHODS: A year-long fellowship was created for junior URM academic clinician faculty with funding through the Society of Teachers of Family Medicine Project Fund. Seven junior faculty applied and were accepted to participate in the fellowship, which included conference calls and an in-person workshop covering topics related to writing and career advancement. RESULTS: The workshop included a mix of prepared programming on how to move from idea to project to manuscript, as well as time for spontaneous mentorship and manuscript collaboration. Key themes that emerged included how to address the high cost of the minority tax, the need for individual passion as a pathway to success, and how to overcome imposter syndrome as a hindrance to writing. CONCLUSIONS: The "for URM by URM" approach for faculty development to promote writing skills and scholarship for junior URM Family Medicine physicians can address challenges faced by URM faculty. By using a framework that includes the mentors' lived experiences and creates a psychological safe space, we can address concerns often overlooked in traditional skills-based faculty development programs.

Group Prenatal Care and Maternal Outcomes: A Scoping Review.

PURPOSE: The objective of this study was to examine the current state of literature on group prenatal care and its impact on maternal outcomes and racial disparities in adverse maternal outcomes. DESIGN: We conducted a scoping review of literature published between January 2010 and December 2020 using the PRISMA-ScR reporting checklist. METHODS: Eligible studies were identified using key words and MeSH terms in PubMed, CINAHL, and Web of Science. Inclusion criteria were studies that were (a) conducted in the United States; (b) published between January 2010 and December 2020; (c) in English; (d) focused on the primary investigation of group prenatal care and reporting on maternal comorbidity outcomes; and (e) an observational study or clinical trial. RESULTS: Nine studies met inclusion criteria. They reported on outcomes of preeclampsia, gestational hypertension, gestational diabetes mellitus, final A1C among patients with gestational diabetes mellitus, and postpartum hemorrhage. None reported on racial disparities for minoritized populations. Among all reported maternal outcomes, results were mixed, providing inconclusive evidence. CLINICAL IMPLICATIONS: Outcomes from group prenatal care focus more on neonatal outcomes than maternal outcomes. More studies are needed with stronger designs. Given pervasive racial disparities in U.S. maternal mortality, future studies should assess how group prenatal care participation may contribute to fewer experiences of racial discrimination and implicit bias for Black women in maternity care.

Medial femoral trochlea flap reconstruction versus proximal row carpectomy for Kienböck's disease: a morphometric comparison.

Surgical options for advanced Kienböck's disease include proximal row carpectomy or lunate reconstruction with a medial femoral trochlea osteochondral flap. This study compares morphology of the proximal capitate and the medial femoral trochlear surfaces to the proximal lunate using three-dimensional geometric morphometric analysis. Virtual articular surfaces were extracted from MRI studies of ten healthy volunteers. Distances between corresponding points on the proximal lunate and proximal capitate or medial femoral trochlear surfaces were measured. In seven subjects, mean inter-surface distance for the medial femoral trochlea-proximal lunate pair was significantly lower than the proximal capitate-proximal lunate pairing. In three subjects, mean proximal capitate-proximal lunate distance was significantly lower. We conclude that the medial femoral trochlear flap was anatomically closer to the shape of the proximal lunate in the majority of the examined subjects. However, we found that in three out of ten cases, the proximal capitate was a better match.

Use of an adeno-associated virus serotype Anc80 to provide durable cure of phenylketonuria in a mouse model.

Phenylketonuria (PKU) is the most common inborn error of metabolism of the liver, and results from mutations of both alleles of the phenylalanine hydroxylase gene (PAH). As such, it is a suitable target for gene therapy via gene delivery with a recombinant adeno-associated virus (AAV) vector. Here we use the synthetic AAV vector Anc80 via systemic administration to deliver a functional copy of a codon-optimized human PAH gene, with or without an intron spacer, to the Pahenu2 mouse model of PKU. Dose-dependent transduction of the liver and expression of PAH mRNA were present with both vectors, resulting in significant and durable reduction of circulating phenylalanine, reaching near control levels in males. Coat color of treated Pahenu2 mice reflected an increase in pigmentation from brown to the black color of control animals, further indicating functional restoration of phenylalanine metabolism and its byproduct melanin. There were no adverse effects associated with administration of AAV up to 5 × 1012 VG/kg, the highest dose tested. Only minor and/or transient variations in some liver enzymes were observed in some of the AAV-dosed animals which were not associated with pathology findings in the liver. Finally, there was no impact on cell turnover or apoptosis as evaluated by Ki-67 and TUNEL staining, further supporting the safety of this approach. This study demonstrates the therapeutic potential of AAV Anc80 to safely and durably cure PKU in a mouse model, supporting development for clinical consideration.

Development of a porcine model of phenylketonuria with a humanized R408W mutation for gene editing.

Phenylketonuria (PKU) is a metabolic disorder whereby phenylalanine metabolism is deficient due to allelic variations in the gene for phenylalanine hydroxylase (PAH). There is no cure for PKU other than orthotopic liver transplantation, and the standard of care for patients is limited to dietary restrictions and key amino acid supplementation. Therefore, Pah was edited in pig fibroblasts for the generation of PKU clone piglets that harbor a common and severe human mutation, R408W. Additionally, the proximal region to the mutation was further humanized by introducing 5 single nucleotide polymorphisms (SNPs) to allow for development of gene editing machinery that could be translated directly from the pig model to human PKU patients that harbor at least one classic R408W allele. Resulting piglets were hypopigmented (a single Ossabaw piglet) and had low birthweight (all piglets). The piglets had similar levels of PAH expression, but no detectable enzymatic activity, consistent with the human phenotype. The piglets were fragile and required extensive neonatal care to prevent failure to thrive and early demise. Phenylalanine levels rose sharply when dietary Phe was unrestricted but could be rapidly reduced with a low Phe diet. Fibroblasts isolated from R408W piglets show susceptibility to correction using CRISPR or TALEN, with subsequent homology-directed recombination to correct Pah. This pig model of PKU provides a powerful new tool for development of all classes of therapeutic candidates to treat or cure PKU, as well as unique value for proof-of-concept studies for in vivo human gene editing platforms in the context of this humanized PKU allele.

Ex Vivo Cell Therapy by Ectopic Hepatocyte Transplantation Treats the Porcine Tyrosinemia Model of Acute Liver Failure.

The effectiveness of cell-based therapies to treat liver failure is often limited by the diseased liver environment. Here, we provide preclinical proof of concept for hepatocyte transplantation into lymph nodes as a cure for liver failure in a large-animal model with hereditary tyrosinemia type 1 (HT1), a metabolic liver disease caused by deficiency of fumarylacetoacetate hydrolase (FAH) enzyme. Autologous porcine hepatocytes were transduced ex vivo with a lentiviral vector carrying the pig Fah gene and transplanted into mesenteric lymph nodes. Hepatocytes showed early (6 h) and durable (8 months) engraftment in lymph nodes, with reproduction of vascular and hepatic microarchitecture. Subsequently, hepatocytes migrated to and repopulated the native diseased liver. The corrected cells generated sufficient liver mass to clinically ameliorate the acute liver failure and HT1 disease as early as 97 days post-transplantation. Integration site analysis defined the corrected hepatocytes in the liver as a subpopulation of hepatocytes from lymph nodes, indicating that the lymph nodes served as a source for healthy hepatocytes to repopulate a diseased liver. Therefore, ectopic transplantation of healthy hepatocytes cures this pig model of liver failure and presents a promising approach for the development of cures for liver disease in patients.

Comparison of Gene Delivery to the Kidney by Adenovirus, Adeno-Associated Virus, and Lentiviral Vectors After Intravenous and Direct Kidney Injections.

There are many kidney diseases that might be addressed by gene therapy. However, gene delivery to kidney cells is inefficient. This is due, in part, to the fact that the kidney excludes molecules above 50 kDa and that most gene delivery vectors are megaDaltons in mass. We compared the ability of adeno-associated virus (AAV), adenovirus (Ad), and lentiviral (LV) vectors to deliver genes to renal cells. When vectors were delivered by the intravenous (IV) route in mice, weak luciferase activity was observed in the kidney with substantially more in the liver. When gene delivery was observed in the kidney, expression was primarily in the glomerulus. To avoid these limitations, vectors were injected directly into the kidney by retrograde ureteral (RU) and subcapsular (SC) injections in mice. Small AAV vectors transduced the kidney, but also leaked from the organ and mediated higher levels of transduction in off-target tissues. Comparison of AAV2, 6.2, 8, and rh10 vectors by direct kidney injection demonstrated highest delivery by AAV6.2 and 8. Larger Ad and LV vectors transduced kidney cells and mediated less off-target tissue transduction. These data demonstrate the utility of direct kidney injections to circumvent the kidney size exclusion barrier. They also identify the effects of vector size on on-target and off-target transduction. This lays the foundation for the use of different vector platforms for gene therapy of diverse kidney diseases.

Ex Vivo Hepatocyte Reprograming Promotes Homology-Directed DNA Repair to Correct Metabolic Disease in Mice After Transplantation.

Ex vivo CRISPR/Cas9-mediated gene editing in hepatocytes using homology-directed repair (HDR) is a potential alternative curative therapy to organ transplantation for metabolic liver disease. However, a major limitation of this approach in quiescent adult primary hepatocytes is that nonhomologous end-joining is the predominant DNA repair pathway for double-strand breaks (DSBs). This study explored the hypothesis that ex vivo hepatocyte culture could reprogram hepatocytes to favor HDR after CRISPR/Cas9-mediated DNA DSBs. Quantitative PCR (qPCR), RNA sequencing, and flow cytometry demonstrated that within 24 hours, primary mouse hepatocytes in ex vivo monolayer culture decreased metabolic functions and increased expression of genes related to mitosis progression and HDR. Despite the down-regulation of hepatocyte function genes, hepatocytes cultured for up to 72 hours could robustly engraft in vivo. To assess functionality long-term, primary hepatocytes from a mouse model of hereditary tyrosinemia type 1 bearing a single-point mutation were transduced ex vivo with two adeno-associated viral vectors to deliver the Cas9 nuclease, target guide RNAs, and a 1.2-kb homology template. Adeno-associated viral Cas9 induced robust cutting at the target locus, and, after delivery of the repair template, precise correction of the point mutation occurred by HDR. Edited hepatocytes were transplanted into recipient fumarylacetoacetate hydrolase knockout mice, resulting in engraftment, robust proliferation, and prevention of liver failure. Weight gain and biochemical assessment revealed normalization of metabolic function. Conclusion: The results of this study demonstrate the potential therapeutic effect of ex vivo hepatocyte-directed gene editing after reprogramming to cure metabolic disease in a preclinical model of hereditary tyrosinemia type 1.

Hepatotoxicity and Toxicology of In Vivo Lentiviral Vector Administration in Healthy and Liver-Injury Mouse Models.

General safety and toxicology assessments supporting in vivo lentiviral vector-based therapeutic development are sparse. We have previously demonstrated the efficacy of a lentiviral vector expressing fumarylacetoacetate hydrolase (LV-FAH) to cure animal models of hereditary tyrosinemia type 1. Therefore, we performed a complete preclinical toxicological evaluation of LV-FAH, in a large cohort (n = 20/group) of wildtype mice and included matched groups of N-nitrosodiethylamine/carbon tetrachloride (DEN/CCl4)-induced liver injury mice to assess specific toxicity in fibrotic liver tissue. Mice receiving LV-FAH alone (109 TU/mouse) or in combination with DEN/CCl4 presented clinically similar to control animals, with only slight reductions in total body weight gains over the study period (3.2- to 3.7-fold vs. 4.2-fold). There were no indications of toxicity attributed to administration of LV-FAH alone over the duration of this study. The known hepatotoxic combination of DEN/CCl4 induced fibrotic liver injury, and co-administration with LV-FAH was associated with exaggeration of some findings such as an increased liver:body weight ratio and progression to focal hepatocyte necrosis in some animals. Hepatocellular degeneration/regeneration was present in DEN/CCl4-dosed animals regardless of LV-FAH as evaluated by Ki-67 immunohistochemistry and circulating alpha fetoprotein levels, but there were no tumors identified in any tissue in any dose group. These data demonstrate the inherent safety of LV-FAH and support broader clinical development of lentiviral vectors for in vivo administration.

Autologous Gene and Cell Therapy Provides Safe and Long-Term Curative Therapy in A Large Pig Model of Hereditary Tyrosinemia Type 1.

Orthotopic liver transplantation remains the only curative therapy for inborn errors of metabolism. Given the tremendous success for primary immunodeficiencies using ex-vivo gene therapy with lentiviral vectors, there is great interest in developing similar curative therapies for metabolic liver diseases. We have previously generated a pig model of hereditary tyrosinemia type 1 (HT1), an autosomal recessive disorder caused by deficiency of fumarylacetoacetate hydrolase (FAH). Using this model, we have demonstrated curative ex-vivo gene and cell therapy using a lentiviral vector to express FAH in autologous hepatocytes. To further evaluate the long-term clinical outcomes of this therapeutic approach, we continued to monitor one of these pigs over the course of three years. The animal continued to thrive off the protective drug NTBC, gaining weight appropriately, and maintaining sexual fecundity for the course of his life. The animal was euthanized 31 months after transplantation to perform a thorough biochemical and histological analysis. Biochemically, liver enzymes and alpha-fetoprotein levels remained normal and abhorrent metabolites specific to HT1 remained corrected. Liver histology showed no evidence of tumorigenicity and Masson's trichrome staining revealed minimal fibrosis and no evidence of cirrhosis. FAH-immunohistochemistry revealed complete repopulation of the liver by transplanted FAH-positive cells. A complete histopathological report on other organs, including kidney, revealed no abnormalities. This study is the first to demonstrate long-term safety and efficacy of hepatocyte-directed gene therapy in a large animal model. We conclude that hepatocyte-directed ex-vivo gene therapy is a rational choice for further exploration as an alternative therapeutic approach to whole organ transplantation for metabolic liver disease, including HT1.

Lentiviral Vector-mediated Gene Therapy of Hepatocytes Ex Vivo for Autologous Transplantation in Swine.

Gene therapy is an ideal choice to cure many inborn errors of metabolism of the liver. Ex-vivo, lentiviral vectors have been used successfully in the treatment of many hematopoietic diseases in humans, as their use offers stable transgene expression due to the vector's ability to integrate into the host genome. This method demonstrates the application of ex vivo gene therapy of hepatocytes to a large animal model of hereditary tyrosinemia type I. This process consists of 1) isolation of primary hepatocytes from the autologous donor/recipient animal, 2) ex vivo gene delivery via hepatocyte transduction with a lentiviral vector, and 3) autologous transplant of corrected hepatocytes via portal vein injection. Success of the method generally relies upon efficient and sterile removal of the liver resection, careful handling of the excised specimen for isolation of viable hepatocytes sufficient for re-engrafting, high-percentage transduction of the isolated cells, and aseptic surgical procedures throughout to prevent infection. Technical failure at any of these steps will result in low yield of viable transduced hepatocytes for autologous transplant or infection of the donor/recipient animal. The pig model of human type 1 hereditary tyrosinemia (HT-1) chosen for this approach is uniquely amenable to such a method, as even a small percentage of engraftment of corrected cells will lead to repopulation of the liver with healthy cells based on a powerful selective advantage over native-diseased hepatocytes. Although this growth selection will not be true for all indications, this approach is a foundation for expansion into other indications and allows for manipulation of this environment to address additional diseases, both within the liver and beyond, while controlling for exposure to viral vector and opportunity for off-target toxicity and tumorigenicity.