Skin in the game: a review of single-cell and spatial transcriptomics in dermatological research.

Single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics (ST) are two emerging research technologies that uniquely characterize gene expression microenvironments on a cellular or subcellular level. The skin, a clinically accessible tissue composed of diverse, essential cell populations, serves as an ideal target for these high-resolution investigative approaches. Using these tools, researchers are assembling a compendium of data and discoveries in healthy skin as well as a range of dermatologic pathophysiologies, including atopic dermatitis, psoriasis, and cutaneous malignancies. The ongoing advancement of single-cell approaches, coupled with anticipated decreases in cost with increased adoption, will reshape dermatologic research, profoundly influencing disease characterization, prognosis, and ultimately clinical practice.

Lipid Nanoparticles Delivering Constitutively Active STING mRNA to Stimulate Antitumor Immunity.

Treating immunosuppressive tumors represents a major challenge in cancer therapies. Activation of STING signaling has shown remarkable potential to invigorate the immunologically "cold" tumor microenvironment (TME). However, we have shown that STING is silenced in many human cancers, including pancreatic ductal adenocarcinoma (PDAC) and Merkel cell carcinoma (MCC). In this study, we demonstrated that mRNA-lipid nanoparticle (LNP) technology could be used to efficiently deliver naturally occurring constitutively active STING mutant STINGR284S into these cancer cells to reactivate STING antitumor immunity and trigger robust killing of tumor cells. STING agonists are being actively pursued as cancer immunotherapies. However, traditional STING agonists can induce T cell cytotoxicity, counteracting the desired antitumor immune response. In addition, the antitumor efficacy of traditional STING agonists obligatorily depends on STING expression and does not work in STING-silenced cancers. Importantly, we found that STINGR284S mRNA-LNP does not introduce T cell cytotoxicity. Our studies demonstrated that mRNA-LNP delivery of STINGR284S can reactivate the antitumor response without introducing antiproliferative effects in lymphocytic immune cells, overcoming the toxicity and limitations of conventional STING agonists. Our work therefore identifies a novel therapeutic tool for reactivating antitumor immunity in an array of STING-silenced immunologically "cold" tumors that are refractory to current therapies.

A 3-Dimensional Biomimetic Platform to Interrogate the Safety of Autologous Fat Transfer in the Setting of Breast Cancer.

INTRODUCTION: Obesity is a known risk factor for the development and prognosis of breast cancer. Adipocytes have been identified as a source of exogenous lipids in other cancer types and may similarly provide energy to fuel malignant survival and growth in breast cancer. This relationship is of particular relevance to plastic surgery, because many reconstructions after oncologic mastectomy achieve optimal aesthetics and durability using adjunctive autologous fat transfer (AFT). Despite the increasing ubiquity and promise of AFT, many unanswered questions remain, including safety in the setting of breast cancer. Clinical studies to examine this question are underway, but an in vitro system is critical to elucidate the complex interplay between the cells that normally reside at the surgical recipient site. To study these interactions and characterize possible lipid transfer between adipocytes to breast cancer cells, we designed a 3-dimensional in vitro model using primary patient-derived tissues. METHODS: Breast adipose tissue was acquired from patients undergoing breast reduction surgery. The tissue was enzymatically digested and sorted to retrieve adipocytes and adipose stromal cells. Polydimethylsiloxane wells were filled with type I collagen-encapsulated adipocytes labeled with the fluorescent lipid dye boron dipyrromethene, as well as unlabeled adipose stromal cells. A monolayer of red fluorescently labeled MDA-MB-231 and MDA-MB-468 breast cancer cells was seeded on the surface of the construct. Lipid transfer at the interface between adipocytes and breast cancer cells was analyzed. RESULTS: Confocal microscopy revealed a dense culture of native adipocytes containing fluorescent lipid droplets in the 3-dimensional collagen culture platform. RFP-positive breast cancer cells were found in close proximity to lipid-laden adipocytes. Lipid transfer from adipocytes to breast cancer cells was observed by the presence of boron dipyrromethene-positive lipid droplets within RFP-labeled breast cancer cells. CONCLUSION: We have established a 3-dimensional model to study complex breast cancer-adipose tissue interactions. Direct transfer of fluorescently labeled lipids from adipocytes to breast cancer cells may indicate aberrant metabolism to fuel malignant growth and adaptive survival. Our novel platform can untangle the complex interplay within the breast cancer tumor microenvironment for high-throughput analysis and better elucidate the safety of AFT in postoncologic mastectomy.

Characterization of assembly intermediates containing subunit 1 of yeast cytochrome oxidase.

Mitochondrial-encoded Cox1p, one of the three core subunits of yeast cytochrome oxidase (COX), was previously shown to associate with regulatory proteins and nuclear-encoded subunits into five high molecular weight complexes that were proposed to constitute the pathway for biogenesis of the Cox1p assembly module. One of the intermediates (D5) was inferred, but not directly shown to exist. In the present study mitochondria of strains expressing C-terminal-tagged subunits of COX that had not been looked at previously were pulse-labeled and analyzed for the presence of newly translated Cox1p in the immunoprecipitates. These studies revealed that of the eight nuclear-encoded COX subunits, only Cox5ap, Cox6p, and Cox8p are present in the Cox1p module. Both Cox5ap and Cox8p share interfaces with Cox1p in the holoenzyme, whereas Cox6p interacts indirectly through Cox5ap. These results suggest that the subunit contacts in the holoenzyme are probably established during biogenesis of the Cox1p module. To confirm the existence of the largest Cox1p intermediates (D5), which was only inferred previously, radiolabeled Cox1p with a C-terminal tag was expressed in COX-deficient pet111 and pet494 mutants. Pulldown assays confirmed the presence of newly translated Cox1p in D5, which in wild type cannot be demonstrated directly because of its co-migration with COX in the native electrophoresis system used to separate the intermediates. Jointly, the results of these analyses substantiate our previous proposal that COX is assembled from separate assembly modules, each containing one of the mitochondrial-translated core subunits in association with a unique set of nuclear-encoded subunits.

Epicanthoplasty: Social and historical perspectives.

The epicanthus is a fold of skin covering the inner corner of the eye which blends into the nasal skin. It is a cosmetic feature of many populations of the world. The surgical alteration of this structure was first developed for the epicanthus found in such congenital genetic conditions as Down syndrome in the West. In the past century and a half, in what may be a reaction to the Western portrayal of skin overlying the eye and of Shakespeare's descriptions of characters with epicanthic folds, surgical techniques have arisen for pure cosmetic intent to alter the Asian eyelid. These procedures have almost wholly become undertaken by patients and surgeons of East Asian descent. Since 1989, the epicanthus surgery literature has been penned predominantly by authors of East Asian descent.

Parallel lives in contrast: Scars in the Roman Republic versus English hereditary monarchy.

Scars are a visible part of the political forum in the Roman Republic and in English hereditary monarchy. Coriolanus's scars are celebrated by Romans in William Shakespeare's Coriolanus; in contrast, an absent record of King Richard II's skin ever breaking is part of the collective fiction of hereditary monarchy in Shakespeare's Richard II. For democracy in Rome, the symbology of scarring may be a practical element in ratifying the office of the consul: as a reminder of Rome's experience with the Tarquin kings they had expelled and to avoid the concentration of power in any one man. Consuls would serve only one year, and there were two consuls each year. Scars demonstrated that these politicians were not gods like kings wished to be. In parallel, scar-free, impenetrable skin preserves King Richard II's symbolism as an anointed monarch. Henry Bolingbroke and his son, Prince Hal and future Henry V, face the psychologic and political consequences of stripping the kingship of this aura, by demonstrating that a king's skin can be penetrated on the battlefield by any of his subjects.

The royal touch: Scrofula and defining monarchy.

English monarchs have claimed the ability to heal scrofula since the turn of the first millennium, long before the mycobacterial cause was identified in the late 19th century. A disease with a self-limiting clinical course, scrofula helped to define a period of human political organization in Western Europe known as monarchy. Termed the royal touch, the practice of touching subjects and curing them of scrofula demonstrated that a monarch belonged to the legitimate dynasty. Despite this, James VI & I (1566-1625) was unenthusiastic about touching his subjects, when he ostensibly had the most to gain from its symbolism; he was fanatical about the absolute political power of monarchs. Shakespeare's (1564-1616) inclusion of the royal touch in Macbeth, written as King James I ascended the English throne after Queen Elizabeth I's (1533-1603) death, may be a subtle nod to the outsider Scottish king to recognize the political value of the tradition. In contrast to King James, his descendants intended to reinforce their affiliation with the royal touch, adapt the royal touch for a modern constitutional monarchy, and memorialize how those in their dynasty touched the skin of their most disfigured subjects.

Salt-inducible kinase inhibition promotes the adipocyte thermogenic program and adipose tissue browning.

OBJECTIVE: Norepinephrine stimulates the adipose tissue thermogenic program through a ß-adrenergic receptor (ßAR)-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signaling cascade. We discovered that a noncanonical activation of the mechanistic target of rapamycin complex 1 (mTORC1) by PKA is required for the ßAR-stimulation of adipose tissue browning. However, the downstream events triggered by PKA-phosphorylated mTORC1 activation that drive this thermogenic response are not well understood. METHODS: We used a proteomic approach of Stable Isotope Labeling by/with Amino acids in Cell culture (SILAC) to characterize the global protein phosphorylation profile in brown adipocytes treated with the ßAR agonist. We identified salt-inducible kinase 3 (SIK3) as a candidate mTORC1 substrate and further tested the effect of SIK3 deficiency or SIK inhibition on the thermogenic gene expression program in brown adipocytes and in mouse adipose tissue. RESULTS: SIK3 interacts with RAPTOR, the defining component of the mTORC1 complex, and is phosphorylated at Ser884 in a rapamycin-sensitive manner. Pharmacological SIK inhibition by a pan-SIK inhibitor (HG-9-91-01) in brown adipocytes increases basal Ucp1 gene expression and restores its expression upon blockade of either mTORC1 or PKA. Short-hairpin RNA (shRNA) knockdown of Sik3 augments, while overexpression of SIK3 suppresses, Ucp1 gene expression in brown adipocytes. The regulatory PKA phosphorylation domain of SIK3 is essential for its inhibition. CRISPR-mediated Sik3 deletion in brown adipocytes increases type IIa histone deacetylase (HDAC) activity and enhances the expression of genes involved in thermogenesis such as Ucp1, Pgc1α, and mitochondrial OXPHOS complex protein. We further show that HDAC4 interacts with PGC1α after ßAR stimulation and reduces lysine acetylation in PGC1α. Finally, a SIK inhibitor well-tolerated in vivo (YKL-05-099) can stimulate the expression of thermogenesis-related genes and browning of mouse subcutaneous adipose tissue. CONCLUSIONS: Taken together, our data reveal that SIK3, with the possible contribution of other SIKs, functions as a phosphorylation switch for ß-adrenergic activation to drive the adipose tissue thermogenic program and indicates that more work to understand the role of the SIKs is warranted. Our findings also suggest that maneuvers targeting SIKs could be beneficial for obesity and related cardiometabolic disease.

Neutralization against Omicron subvariants after BA.5/BF.7 breakthrough infection weakened as virus evolution and aging despite repeated prototype-based vaccination1.

BACKGROUND: Omicron had swept the mainland China between December 2022 and January 2023, while SARS-CoV-2 still continued to evolve. To fully prepare for the next wave, it's urgent to evaluate the humoral immune response post BA.5/BF.7 breakthrough infection against predominant sub-lineages among existing vaccination strategies and the elders. METHOD: This study enrolled a longitudinal young-adult cohort from 2/3-dose vaccination to 1 month after breakthrough infection, and an elder cohort at 1 month after breakthrough infection. Seral samples were collected and tested for humoral immune response to SARS-CoV-2 subvariants including WT, BA.2, BA.5, BF.7, BQ.1.1, CH.1.1, XBB.1.5. RESULTS: BA.5/BF.7 breakthrough infection induced higher neutralization activity than solely vaccination in all SARS-CoV-2 strains, while the latest Omicron subvariants, BQ.1.1, CH.1.1, XBB.1.5, exhibited the strongest neutralization evasion ability. There was a negative correlation between age and humoral immune response in WT, BA.5, BQ.1.1, and XBB.1.5. Compared to non-vaccination groups, breakthrough infection in two-dose vaccination groups had significantly higher neutralizing antibody against WT, BA.2, BA.5, BF.7 but not to BQ.1.1, CH.1.1, XBB.1.5 while booster dose against the prototype prior-breakthrough would not further significantly enhance individual's humoral responses against the latest Omicron subvariants. CONCLUSIONS: Newer variants manifest increasing immune evasion from neutralization and repeated prototype-based booster vaccines may not further enhance neutralizing antibody against emerging new variants. Older adults have lower levels of neutralizing antibody. Future vaccination strategies should aim to enhance effective neutralization to contemporary variants.

Seminal vesicle mass: An unusual site for metastatic hepatocellular carcinoma after orthotopic liver transplant.

Metastasis of hepatocellular carcinoma (HCC) to the seminal vesicle is extraordinarily rare, with only two other cases reported in the literature. Herein we present the first documented case of a seminal vesicle as the initial site of solitary metastasis in a patient with a history of liver transplantation for HCC. We aim to provide more information regarding the disease process, histopathology, and management strategy.

On Shakespeare and cutaneous diseases.

Disorders with dermatologic features are intractable and rife in Shakespeare's world. For this reason, they occupy an unusual position in culture. "The plague" and "leprosy" are popular insults and epithets-indictments of not external pathogens but of moral failure. It may be no surprise to identify, as a present-day reader, syphilis and the plague in early modern England, but what about other dermatologic conditions at the time? "The plague," "leprosy," "the pox," and "measles" are commonly used terms, although ultimately interchangeable in Shakespeare's plays. Rosacea and scurvy, however, are described by Shakespeare before they become named entities. Bardolph's skin characterizes his alcohol addiction and thievery, and Caliban's "monstrous" form mirrors his state as an untouchable in society. Shakespeare also documents the "royal touch" in Macbeth, which links the ability of a monarch to heal the skin lesions of extrapulmonary tuberculosisto political legitimacy. Shakespeare does little more than document this historic ritual around the skin, but he does catalogue the ways these acquired diseases of the skin are used in daily speech and character writing. At the end of Macbeth, the "royal touch" gives Prince Malcolm the backstory and political precedent to march on Dunsinane Hill to reclaim the Scottish throne.

Current Perspectives on the Diagnosis and Management of Primary Urethral Cancer: A Systematic Review.

Primary urethral cancer (PUC) is a rare but highly aggressive malignancy that causes malignant urethral obstruction. We conducted a literature review using PubMed to identify original research studies that assessed the diagnosis and management of primary urethral cancer. PUC affects men more than women, is more common in African Americans than Caucasians, and is associated with history of chronic inflammation and irritation of the urinary tract. Patients suspected of PUC should undergo a complete work-up including cystoscopy, magnetic resonance imaging, and biopsy. In men and women, surgical monotherapy ranging from organ-sparing to more radical reconstructive procedures has adequate survival rates for early stage PUC and has been shown to be similarly as effective as radiation monotherapy, while multimodal therapy has become the standard of treatment for advanced stage PUC. Salvage surgery or radiation therapy has been linked with increased survival rates. Nodal involvement at the time of diagnosis is a negative prognosticator and should be treated with multimodal therapy. Further prospective studies with greater sample sizes and standardized clinical trials would allow for greater consistency in evaluating the different treatment modalities for PUC.

On the diagnostic and neurobiological origins of bipolar disorder.

Psychiatry is constructed around a taxonomy of several hundred diagnoses differentiated by nuances in the timing, co-occurrence, and severity of symptoms. Bipolar disorder (BD) is notable among these diagnoses for manic, depressive, and psychotic symptoms all being core features. Here, we trace current understanding of the neurobiological origins of BD and related diagnoses. To provide context, we begin by exploring the historical origins of psychiatric taxonomy. We then illustrate how key discoveries in pharmacology and neuroscience gave rise to a generation of neurobiological hypotheses about the origins of these disorders that facilitated therapeutic innovation but failed to explain disease pathogenesis. Lastly, we examine the extent to which genetics has succeeded in filling this void and contributing to the construction of an objective classification of psychiatric disturbance.

Regulation of mitochondrial translation of the ATP8/ATP6 mRNA by Smt1p.

Expression of the mitochondrially encoded ATP6 and ATP8 genes is translationally regulated by F1 ATPase. We report a translational repressor (Smt1p) of the ATP6/8 mRNA that, when mutated, restores translation of the encoded Atp6p and Atp8p subunits of the ATP synthase. Heterozygous smt1 mutants fail to rescue the translation defect, indicating that the mutations are recessive. Smt1p is an intrinsic inner membrane protein, which, based on its sedimentation, has a native size twice that of the monomer. Affinity purification of tagged Smt1p followed by reverse transcription of the associated RNA and PCR amplification of the resultant cDNA with gene-specific primers demonstrated the presence in mitochondria of Smt1p-ATP8/ATP6 and Smt1p-COB mRNA complexes. These results indicate that Smt1p is likely to be involved in translational regulation of both mRNAs. Applying Occam's principle, we favor a mechanistic model in which translation of the ATP8/ATP6 bicistronic mRNA is coupled to the availability of F1 for subsequent assembly of the Atp6p and Atp8p products into the ATP synthase. The mechanism of this regulatory pathway is proposed to entail a displacement of the repressor from the translationally mute Smt1-ATP8/ATP6 complex by F1, thereby permitting the Atp22p activator to interact with and promote translation of the mRNA.