Cardiometabolic outcomes in Kronos Early Estrogen Prevention Study continuation: 14-year follow-up of a hormone therapy trial.

OBJECTIVE: This study aimed to determine long-term cardiometabolic effects of hormone therapies initiated within 3 years of onset of menopause after a 14-year follow-up study of participants of the Kronos Early Estrogen Prevention Study (KEEPS). METHODS: KEEPS was a multisite clinical trial that recruited recently menopausal women with good cardiovascular health for randomization to oral conjugated equine estrogens (Premarin, 0.45 mg/d) or transdermal 17ß-estradiol (Climara, 50 µg/d) both with micronized progesterone (Prometrium, 200 mg/d) for 12 d/mo, or placebo pills and patch for 4 years. KEEPS continuation recontacted KEEPS participants 14 years after randomization and 10 years after the completion of the 4-year clinical trial to attend in-person clinic visits. RESULTS: Participants of KEEPS continuation (n = 299 of the 727 KEEPS participants; 41%) had an average age of 67 years (range, 58-73 y). Measurements of systolic and diastolic blood pressures, waist-to-hip ratio, fasting levels of glucose, insulin, lipid profiles, and homeostasis model assessment of insulin resistance were not different among the treatment groups at either KEEPS baseline or at KEEPS continuation visits, or for change between these two visits. The frequency of self-reported diabetes ( P = 0.007) and use of diabetes medications was higher in the placebo than the oral conjugated equine estrogens ( P = 0.045) or transdermal 17ß-estradiol ( P = 0.02) groups, but these differences were not supported by the laboratory measurements of glycemia or insulin resistance. CONCLUSIONS: There was no evidence of cardiovascular and/or metabolic benefits or adverse effects associated with 4 years use of oral or transdermal forms of hormone therapy by recently menopausal women with good cardiovascular health after 10 years.

Machine learning modeling practices to support the principles of AI and ethics in nutrition research.

BACKGROUND: Nutrition research is relying more on artificial intelligence and machine learning models to understand, diagnose, predict, and explain data. While artificial intelligence and machine learning models provide powerful modeling tools, failure to use careful and well-thought-out modeling processes can lead to misleading conclusions and concerns surrounding ethics and bias. METHODS: Based on our experience as reviewers and journal editors in nutrition and obesity, we identified the most frequently omitted best practices from statistical modeling and how these same practices extend to machine learning models. We next addressed areas required for implementation of machine learning that are not included in commercial software packages. RESULTS: Here, we provide a tutorial on best artificial intelligence and machine learning modeling practices that can reduce potential ethical problems with a checklist and guiding principles to aid nutrition researchers in developing, evaluating, and implementing artificial intelligence and machine learning models in nutrition research. CONCLUSION: The quality of AI/ML modeling in nutrition research requires iterative and tailored processes to mitigate against potential ethical problems or to predict conclusions that are free of bias.

Standard Operating Procedures for Biospecimen Collection, Processing, and Storage: From the Type 1 Diabetes in Acute Pancreatitis Consortium.

ABSTRACT: Differences in methods for biospecimen collection, processing, and storage can yield considerable variability and error. Therefore, best practices for standard operating procedures are critical for successful discovery, development, and validation of disease biomarkers. Here, we describe standard operating procedures developed for biospecimen collection during the DREAM (Diabetes RElated to Acute pancreatitis and its Mechanisms) Study within the Type 1 Diabetes in Acute Pancreatitis Consortium. Notably, these protocols were developed using an integrative process based on prior consortium experience and with input from working groups with expertise in immunology, pancreatitis, and diabetes. Publication and adoption consistent biospecimen protocols will inform future studies and allow for better comparisons across different metabolic research efforts.

Rationale and Development of a Data Coordinating Center to Support the Type 1 Diabetes in Acute Pancreatitis Consortium.

ABSTRACT: A data coordinating center (DCC) is a critical member of any multicenter research undertaking, and that is especially true for the Type 1 Diabetes in Acute Pancreatitis Consortium (T1DAPC). We describe how the T1DAPC DCC supports the consortium via its experience and expertise in project management, administration, financial management, regulatory compliance, scientific coordination, data management, research computing, and biostatistics and in facilitating scientific publications. The DCC's matrix management system has been extremely effective in managing all of its responsibilities. The first 16 months in the life of the T1DAPC have been dedicated to the development of its first protocol, titled Diabetes RElated to Acute pancreatitis and its Mechanisms (DREAM), addressing the institutional review board and regulatory components, developing the T1DAPC data management system, and providing training and certification of clinical center staff. As a result of its efforts, the DCC was a major contributor to the T1DAPC being able to initiate recruitment for the DREAM study in January 2022.

An industrial perspective on co-crystals: Screening, identification and development of the less utilised solid form in drug discovery and development.

Active pharmaceutical ingredients are commonly marketed as a solid form due to ease of transport, storage and administration. In the design of a drug formulation, the selection of the solid form is incredibly important and is traditionally based on what polymorphs, hydrates or salts are available for that compound. Co-crystals, another potential solid form available, are currently not as readily considered as a viable solid form for the development process. Even though co-crystals are gaining an ever-increasing level of interest within the pharmaceutical community, their acceptance and application is still not as standard as other solid forms such as the ubiquitous pharmaceutical salt and stabilised amorphous formulations. Presented in this chapter is information that would allow for a co-crystal screen to be planned and conducted as well as scaled up using solution and mechanochemistry based methods commonly employed in both the literature and industry. Also presented are methods for identifying the formation of a co-crystal using a variety of analytical techniques as well as the importance of confirming the formation of co-crystals from a legal perspective and demonstrating the legal precedent by looking at co-crystalline products already on the market. The benefits of co-crystals have been well established, and presented in this chapter are a selection of examples which best exemplify their potential. The goal of this chapter is to increase the understanding of co-crystals and how they may be successfully exploited in early stage development.

Influence of Ions and pH on the Formation of Solid- and Liquid-like Melanin.

Melanin is a natural pigment with broadband absorption and effective ability to dissipate the energy absorbed. The macromolecular structure of melanin shows a delicate balance between short-range ordered and disordered structures without being a random aggregate. The presence of ions or the variation in pH or ionic strength can alter the self-assembly process which subsequently changes the structure of melanin. To understand these relationships, this study investigates the influence of ions and pH in melanin formation. The types of ions present and pH have a profound influence on the formation and structure of melanin particles, while only minor changes are observed in the absorption and excitation-emission analysis. In some conditions, the formation of discernible particles with significant refractive index contrast is avoided while retaining the spectroscopic characteristics of melanin, leading to liquid-like melanin. These findings identify potential pathways which can be used to manipulate the melanin macromolecular structure while providing the desired spectral properties to enable novel bio-engineering applications.

Self-assembly of ultra-small micelles from amphiphilic lipopeptoids.

Poly(N-substituted glycine) "peptoids" constitute a promising class of peptide-mimetic materials. We introduce the self-assembly of lipopeptoids into spherical micelles ca. 5 nm in diameter as well as larger assemblies by varying the peptoid sequence design. Our results point to design rules for the self-assembly of peptoid nanostructures, enabling the creation of stable, ultra-small peptidomimetic nanospheres.

Patient misconceptions concerning lumbar spondylosis diagnosis and treatment.

OBJECT Patient outcome measures are becoming increasingly important in the evaluation of health care quality and physician performance. Of the many novel measures currently being explored, patient satisfaction and other subjective measures of patient experience are among the most heavily weighted. However, these subjective measures are strongly influenced by a number of factors, including patient demographics, level of understanding of the disorder and its treatment, and patient expectations. In the present study, patients referred to a neurosurgery clinic for degenerative spinal disorders were surveyed to determine their understanding of lumbar spondylosis diagnosis and treatment. METHODS A multiple-choice, 6-question survey was distributed to all patients referred to a general neurosurgical spine clinic at a tertiary care center over a period of 11 months as a quality improvement initiative to assist the provider with individualized patient counseling. The survey consisted of questions designed to assess patient understanding of the role of radiological imaging in the diagnosis and treatment of low-back and leg pain, and patient perception of the indications for surgical compared with conservative management. Demographic data were also collected. RESULTS A total of 121 surveys were included in the analysis. More than 50% of the patients indicated that they would undergo spine surgery based on abnormalities found on MRI, even without symptoms; more than 40% of patients indicated the same for plain radiographs. Similarly, a large proportion of patients (33%) believed that back surgery was more effective than physical therapy in the treatment of back pain without leg pain. Nearly one-fifth of the survey group (17%) also believed that back injections were riskier than back surgery. There were no significant differences in survey responses among patients with a previous history of spine surgery compared with those without previous spine surgery. CONCLUSIONS These results show that a surprisingly high percentage of patients have misconceptions regarding the diagnosis and treatment of lumbar spondylosis, and that these misconceptions persist in patients with a history of spine surgery. Specifically, patients overemphasize the value of radiological studies and have mixed perceptions of the relative risk and effectiveness of surgical intervention compared with more conservative management. These misconceptions have the potential to alter patient expectations and decrease satisfaction, which could negatively impact patient outcomes and subjective valuations of physician performance. While these results are preliminary, they highlight a need for improved communication and patient education during surgical consultation for lumbar spondylosis.

C6-ceramide nanoliposomes target the Warburg effect in chronic lymphocytic leukemia.

Ceramide is a sphingolipid metabolite that induces cancer cell death. When C6-ceramide is encapsulated in a nanoliposome bilayer formulation, cell death is selectively induced in tumor models. However, the mechanism underlying this selectivity is unknown. As most tumors exhibit a preferential switch to glycolysis, as described in the "Warburg effect", we hypothesize that ceramide nanoliposomes selectively target this glycolytic pathway in cancer. We utilize chronic lymphocytic leukemia (CLL) as a cancer model, which has an increased dependency on glycolysis. In CLL cells, we demonstrate that C6-ceramide nanoliposomes, but not control nanoliposomes, induce caspase 3/7-independent necrotic cell death. Nanoliposomal ceramide inhibits both the RNA and protein expression of GAPDH, an enzyme in the glycolytic pathway, which is overexpressed in CLL. To confirm that ceramide targets GAPDH, we demonstrate that downregulation of GAPDH potentiates the decrease in ATP after ceramide treatment and exogenous pyruvate treatment as well as GAPDH overexpression partially rescues ceramide-induced necrosis. Finally, an in vivo murine model of CLL shows that nanoliposomal C6-ceramide treatment elicits tumor regression, concomitant with GAPDH downregulation. We conclude that selective inhibition of the glycolytic pathway in CLL cells with nanoliposomal C6-ceramide could potentially be an effective therapy for leukemia by targeting the Warburg effect.

Depression and pain among inpatients with spinal cord injury and spinal cord disease: differences in symptoms and neurological function.

PURPOSE: To examine the relationship between depression and pain severity during inpatient rehabilitation for those with new onset of spinal cord injury or spinal cord disease (SCI/D), along with patient characteristics, neurological function and etiology. METHOD: This cross-sectional study reviewed medical records of 100 adults with SCI/D including their admission scores on the Patient Health Questionnaire 9 (PHQ-9), a measure of depression, and pain ratings collected at admission and discharge. RESULTS: Upon admission, 28% reported moderate-to-severe symptoms of depression and 69% reported pain. PHQ-9 scores were associated with pain only among those with the least severe impairments, for whom higher scores were associated with greater pain. While depression levels did not differ by etiology, those with traumatic injuries had higher pain ratings. CONCLUSIONS: In general, depressive symptoms were not associated with pain severity in this sample. Etiology was associated with pain, those with traumatic SCI reporting more pain at admission. Among demographic characteristics, age was related to pain, with younger subjects reporting higher levels. These findings suggest that certain characteristics may predispose patients to pain and depression upon admission to rehabilitation. By determining who is at risk for these symptoms, clinicians can adopt treatments that prevent these from becoming chronic conditions.

Seroreactivity to LGL leukemia-specific epitopes in aplastic anemia, myelodysplastic syndrome and paroxysmal nocturnal hemoglobinuria: results of a bone marrow failure consortium study.

Large granular lymphocyte (LGL) leukemia is characterized by clonal expansion of antigen-activated cytotoxic T cells (CTL). Patients frequently exhibit seroreactivity against a human T-cell leukemia virus (HTLV) epitope, BA21. Aplastic anemia, paroxysmal nocturnal hemoglobinuria and myelodysplastic syndrome are bone marrow failure diseases that can also be associated with similar aberrant CTL activation (LGL-BMF). We identified a BA21 peptide that was specifically reactive with LGL leukemia sera and found significantly elevated antibody reactivity against the same peptide in LGL-BMF sera. This finding of shared seroreactivity in LGL-BMF conditions and LGL leukemia suggests that these diseases might share a common pathogenesis.

Therapeutic efficacy of FTY720 in a rat model of NK-cell leukemia.

NK-cell leukemia is a clonal expansion of NK cells. The illness can occur in an aggressive or chronic form. We studied cell lines from human and rat NK-cell leukemias (aggressive NK-cell leukemia) as well as samples from patients with chronic NK-cell leukemia to investigate pathogenic mechanisms. Here we report that Mcl-1 was overexpressed in leukemic NK cells and that knockdown of Mcl-1 induced apoptosis in these leukemic cells. In vitro treatment of human and rat NK leukemia cells with FTY720 led to caspase-dependent apoptosis and decreased Mcl-1 expression in a time- and-dose-dependent manner. These biologic effects could be inhibited by blockade of reactive oxygen species generation and the lysosomal degradation pathway. Lipidomic analyses after FTY720 treatment demonstrated elevated levels of sphingosine, which mediated apoptosis of leukemic NK cells in vitro. Importantly, systemic administration of FTY720 induced complete remission in the syngeneic Fischer rat model of NK-cell leukemia. Therapeutic efficacy was associated with decreased expression of Mcl-1 in vivo. These data demonstrate that therapeutic benefit of FTY720 may result from both altered sphingolipid metabolism as well as enhanced degradation of a key component of survival signaling.

Targeting of survivin by nanoliposomal ceramide induces complete remission in a rat model of NK-LGL leukemia.

The natural killer (NK) type of aggressive large granular lymphocytic (LGL) leukemia is a fatal illness that pursues a rapid clinical course. There are no effective therapies for this illness, and pathogenetic mechanisms remain undefined. Here we report that the survivin was highly expressed in both aggressive and chronic leukemic NK cells but not in normal NK cells. In vitro treatment of human and rat NK-LGL leukemia cells with cell-permeable, short-chain C6-ceramide (C6) in nanoliposomal formulation led to caspase-dependent apoptosis and diminished survivin protein expression, in a time- and dose-dependent manner. Importantly, systemic intravenous delivery of nanoliposomal ceramide induced complete remission in the syngeneic Fischer F344 rat model of aggressive NK-LGL leukemia. Therapeutic efficacy was associated with decreased expression of survivin in vivo. These data suggest that in vivo targeting of survivin through delivery of nanoliposomal C6-ceramide may be a promising therapeutic approach for a fatal leukemia.

Platelet-derived growth factor mediates survival of leukemic large granular lymphocytes via an autocrine regulatory pathway.

Large granular lymphocyte (LGL) leukemia results from chronic expansion of cytotoxic T cells or natural killer (NK) cells. Apoptotic resistance resulting from constitutive activation of survival signaling pathways is a fundamental pathogenic mechanism. Recent network modeling analyses identified platelet-derived growth factor (PDGF) as a key master switch in controlling these survival pathways in T-cell LGL leukemia. Here we show that an autocrine PDGF regulatory loop mediates survival of leukemic LGLs of both T- and NK-cell origin. We found high levels of circulating PDGF-BB in platelet-poor plasma samples from LGL leukemia patients. Production of PDGF-BB by leukemic LGLs was demonstrated by immunocytochemical staining. Leukemic cells expressed much higher levels of PDGFR-beta transcripts than purified normal CD8(+) T cells or NK cells. We observed that phosphatidylinositol-3-kinase (PI3 kinase), Src family kinase (SFK), and downstream protein kinase B (PKB)/AKT pathways were constitutively activated in both T- and NK-LGL leukemia. Pharmacologic blockade of these pathways led to apoptosis of leukemic LGLs. Neutralizing antibody to PDGF-BB inhibited PKB/AKT phosphorylation induced by LGL leukemia sera. These results suggest that targeting of PDGF-BB, a pivotal regulator for the long-term survival of leukemic LGLs, may be an important therapeutic strategy.

MRI-guided dissection of the nonhuman primate brain: a case study.

Numerous biochemical as well as electrophysiological techniques require tissue that must be retrieved very quickly following death in order to preserve the physiological integrity of the neuronal environment. Therefore, the ability to accurately predict the precise locations of brain regions of interest (ROI) and to retrieve those areas as quickly as possible following the brain harvest is critical for subsequent analyses. One way to achieve this objective is the utilization of high-resolution MRI to guide the subsequent dissections. In the present study, individual MRI images of the brains of rhesus and cynomolgus macaques that had chronically self-administered ethanol were employed in order to determine which blocks of dissected tissue contained specific ROIs. MRI-guided brain dissection of discrete brain regions was completely accurate in 100% of the cases. In comparison, approximately 60-70% accuracy was achieved in dissections that relied on external landmarks alone without the aid of MRI. These results clearly demonstrate that the accuracy of targeting specific brain areas can be improved with high-resolution MR imaging.

The role of the prosthetic axilloaxillary loop access as a tertiary arteriovenous access procedure.

PURPOSE: In the last decade, the Dialysis Outcome Quality Initiative (DOQI) Guidelines have enhanced the longevity of patients with end-stage renal disease (ESRD) on hemodialysis. Consequently, surgeons are increasingly challenged to provide vascular access for patients in whom options for access in the upper extremity have been expended. This situation is even more problematic in the morbidly obese patient on hemodialysis. Our group previously reported a high rate of infection and need for secondary interventions in obese patients with prosthetic femorofemoral accesses. We now report a series of patients who underwent placement of a prosthetic axilloaxillary loop access. This study presents our technique and evaluates our results, particularly as they relate to the obese patient. METHODS: From January 1998 to May 2006, 34 prosthetic axilloaxillary loop accesses were placed in 32 patients with ESRD. Eleven patients (12 accesses) were obese, as defined by a body mass index >/=30 kg/m(2). Median follow-up was 16 months. Kaplan-Meier analysis was used to determine primary and secondary patency as well as patient survival for the entire cohort and for the obese and nonobese patient cohorts. Survival curves were compared using the log-rank test for equality over strata. RESULTS: The secondary patency rate was 59% at 1 year (median, 18 months). The 1-year patient survival was 69%. Infection occurred in 15% patients. Comparison of the obese vs nonobese cohorts demonstrated no statistically significant difference in 1-year primary patency (36% vs 10%, P = .17) or secondary patency (71% vs 65%, P = .34). There were no infections in the obese cohort. CONCLUSION: These data show that the prosthetic axilloaxillary loop access has acceptable outcomes and should be considered the tertiary vascular access procedure of choice in the obese patient on hemodialysis.

Pilot study of sex differences in chemokine/cytokine markers of atherosclerosis in humans.

BACKGROUND: Atherogenic processes increase in women after menopause, when the risk of cardiovascular adverse events approaches that observed in age-matched men. In experimental animals, ovariectomy increases the platelet content of mitogenic cytokines, such as platelet-derived growth factor (PDGF), which when released into the blood or site of vascular injury, contribute to atherogenic processes. OBJECTIVE: Experiments were designed to assess the sex distribution of inflammatorychemokines/cytokines, which may be released from platelets in the serum of middle-aged women and men in whom the extent of atherosclerotic coronary disease was defined by coronary arterial calcification (CAC). METHODS: Blood was obtained from healthy white individuals recruited from the Mayo Clinic database. CAC was assessed by 64-slice computed tomography. Plasma cholesterol, lipids, and high-sensitivity C-reactive protein were analyzed by the Mayo Clinic Department of Laboratory Medicine and Pathology. Serum cytokines were determined using cytokine arrays. Cytokine expression was measured using dot blot analysis. RESULTS: Of the 16 individuals (11 women, 5 men) who agreed to participate in the study, 1 woman was premenopausal, 1 was taking oral contraceptives, and 1 was receiving menopausal hormone therapy. One woman had an active infection and was eliminated from the study. CAC was detected in only 2 of the 11 women (scores of 46 and 56 Agatston units [AU]) but in 3 of the 5 men (scores of 3, 123, and 609 AU). Correcting for all other risk factors, expression of the chemokine RANTES (regulated on activation, normal, T-cell expressed and secreted; CCL5 [CC chemokine ligand 5]) was 100.98% greater in women than in men, and PDGF-BB was 55.30% greater in women than in men. CONCLUSIONS: This small pilot study found that the circulating chemokines/cytokines RANTES and PDGF-BB showed sex-disparate distribution between the women and men studied, and did not appear related to the degree of CAC.