Cell and Developmental Biology Commons^™

Junctional Actin As A Mechanical Hub For Supracellular Actomyosin Force Transmission During Apical Constriction, Jasneet Brar

UNLV Theses, Dissertations, Professional Papers, and Capstones

Apical constriction is a conserved morphogenetic mechanism that drives epithelial folding during development. During ventral furrow formation in Drosophila melanogaster, contractile actomyosin networks generate forces that reduce apical cell area resulting in tissue invagination. For these cell-scale forces to produce coordinated tissue-scale changes, contractile networks must be mechanically integrated across neighboring cells. Although adherens junctions are known to mediate this coupling, it is unclear how junctions are integrated into cortical actin cytoskeleton, which directly determines cell, and consequently tissue, shape. This thesis tests the hypothesis that supracellular actomyosin organization during apical constriction depends on distinct adherens junction based mechanical couplings: …

Dysregulation Of Vitamin D Receptor And Cyp450 Metabolism Enzymes May Explain Oral Cancer Responsiveness, Dustin Hunsaker

UNLV Theses, Dissertations, Professional Papers, and Capstones

Many health benefits are associated with Vitamin D (VitD), although deficiency is associated with poor health outcomes and increased risk for cancer development. For example, many tissue specific enzymes are involved in VitD metabolism, and mutations or deletions within Vitamin D receptor (VDR) genes are known to increase cancer risk by altering their functions or bioavailability, although less is known about these phenomena in oral cancers. Using well characterized, commercially available oral cell lines (OKF4, HGF-1, SCC4, SCC9, SCC15, SCC25, CAL27), mRNA expression of P450 cytochrome VitD metabolic enzymes and receptor genes by qPCR revealed differential results. One oral cancer …

Assessing The Effects Of Streptozotocin-Induced Hyperglycemia On Amyloid Beta Accumulation And Cognitive Behavior In A Mouse Model Of Alzheimer’S Disease-Related Pathology, Andrew Adonay Ortiz

UNLV Theses, Dissertations, Professional Papers, and Capstones

Alzheimer’s disease (AD) is an irreversible and devastating neurodegenerative disease characterized by progressive synaptic, dendritic, glial, and neuronal loss, collectively leading to impairments in learning, memory, and deterioration of cognitive and behavioral functions. AD is the most common form of dementia accounting for 60-80% of all cases. AD affects approximately 7 million Americans and is the 6th leading cause of death in the US. Furthermore, by 2050, the US is projected to spend over $1.1 trillion on AD-related treatments. Pathological hallmarks of AD include beta-amyloid (Aβ) plaques, intracellular neurofibrillary tangles (NFTs), and chronic neuroinflammation, which can promote and exacerbate both …

The Present And Future Of Robotic Surgery In Breast Cancer And Breast Reconstruction, Brett Allen, Alexis Knutson, Noama Iftekhar, Casey Giles, Jarrell Patterson, Joshua Macdavid, Richard Baynosa

Department of Plastic Surgery Faculty Research

Background: Breast cancer is the second most common cancer in women with an improving mortality rate and a growing need for reconstruction following oncologic resection. Advancements in robotic surgery and minimally invasive techniques have offered refinement to traditional open techniques of flap harvest for reconstruction, particularly regarding improved donor site morbidity. Methods: The literature review was based on a PubMed database search using the keyword “Robotic breast reconstruction” in conjunction with the Boolean operators “Flap”, “Latissimus”, and “DIEP” to specify the search. In total, 106 results were generated, which were then manually reviewed and condensed for a comprehensive stance on …

Integrated Transcriptomics And Proteomics Study Of Starvation Selected Drosophila Melanogaster, Aavash Adhikari

UNLV Theses, Dissertations, Professional Papers, and Capstones

Drosophila melanogaster, selected for starvation resistance exhibits phenotypic traits such as increased body size, slower development, reduced fecundity and obesity like characteristics including higher lipid reserves and cardiac dysfunction. While prior studies have explored the metabolic and biochemical changes in these flies, the underlying genetic and molecular mechanisms remain unclear. This study aims to investigate the gene expression and protein abundance in starvation-selected Drosophila through RNAseq, proteomics, and integrating both datasets using the MixOmics package in R. I found differential gene expression in the fat body at the third instar larval stage, particularly in pathways related to lipid metabolism, calcium …

Aging-Related Changes In The Mechanical Properties Of Single Cells, Amarnath Singam, Chandrabali Bhattacharya, Seungman Park

Mechanical Engineering Faculty Research

Mechanical properties, along with biochemical and molecular properties, play crucial roles in governing cellular function and homeostasis. Cellular mechanics are influenced by various factors, including physiological and pathological states, making them potential biomarkers for diseases and aging. While several methods such as AFM, particle-tracking microrheology, optical tweezers/stretching, magnetic tweezers/twisting cytometry, microfluidics, and micropipette aspiration have been widely utilized to measure the mechanical properties of single cells, our understanding of how aging affects these properties remains limited. To fill this knowledge gap, we provide a brief overview of the commonly used methods to measure single-cell mechanical properties. We then delve into …

Differential Expression Of Microrna Mir-145 And Mir-155 Downstream Targets In Oral Cancers Exhibiting Limited Chemotherapy Resistance, Conner Belnap

UNLV Theses, Dissertations, Professional Papers, and Capstones

Background: New evidence has suggested that non-coding microRNAs play a significant role in mediating and modulating chemotherapy resistance, particularly among oral cancers. One recent study found that the expression of miR-145 and lack of miR-155 expression strongly correlated with a limited chemotherapy resistance to Cisplatin, 5-Fluorouracil and Paclitaxel , although the mechanism(s) responsible for these observations remain unidentified.

Methods: Using commercially available cell lines of oral squamous cell carcinoma (OSCC), RNA was isolated using phenol:chloroform extraction. The isolated RNA was converted into cDNA by reverse transcription and subsequently screened for the presence or absence of downstream targets of miR-145 and …

Regulation Of Embryonic Eye Regrowth In Xenopus Laevis, Dylan Guerin

UNLV Theses, Dissertations, Professional Papers, and Capstones

Regrowth of lost organs and tissues is an amazing ability that some animals possess. For regrowth to be considered full, the organism must regrow the organ or tissue to a state that is structurally and functionally similar to that which was lost. The ability of an organism to regrow lost or damaged tissue varies among animals, both in the mechanisms utilized to achieve regrowth, and in what body parts can be regrown. Potentially even closely related species can vary wildly in regenerative ability. Therefore, regenerative research can look to other tissues and organisms for direction in elucidating the mechanisms through …

Cellular Mechanisms Of Embryonic Eye Regrowth In Xenopus Laevis, Randolph L. Grell

UNLV Theses, Dissertations, Professional Papers, and Capstones

The African Clawed Frog, Xenopus laevis, has been an indispensable model organism for over 100 years in the study of embryogenesis and development. In recent years, this vertebrate model has emerged as a robust tool for deciphering the cellular and molecular mechanisms underlying various modes of tissue, limb, and organ regeneration. In the context of eye repair, Xenopus offers a unique model, where the adult frog eye has limited regenerative capacity compared to its pre-metamorphic embryonic and tadpole forms, which exhibit a robust capacity for regrowth. Understanding the intricate molecular mechanisms that underlie this developmental shift in regrowth ability …

Treatment Of Warthin’S Tumors Of The Parotid Gland With Radiofrequency Ablation: A Systematic Review Of The Current Literature, Kenny Do, Eric Kawana, Sisi Tian, Jo-Lawrence Bigcas

School of Medicine Faculty Research

Objective: Warthin’s tumors of the parotid gland can be safely observed. Definitive treatment usually requires parotidectomy under general anesthesia. The decision to operate on Warthin’s tumors of the parotid gland can be complicated in patients who wish to avoid risks of surgery and general anesthesia. This systematic review explores the potential of radiofrequency ablation (RFA) as a minimally invasive alternative. Methods: The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) model was used to collect 3 relevant studies that focused on RFA treatment for Warthin’s tumors. The cumulative averages for tumor size and cosmetic scores were then quantified for …

The Use Of Fecal Microbiota Transplant In Overcoming And Modulating Resistance To Anti-Pd-1 Therapy In Patients With Skin Cancer, Tahne Vongsavath, Rodd Rohmani, Kyaw Min Tun, Vignan Manne

Department of Internal Medicine Faculty Research

While immune checkpoint inhibitors have evolved into the standard of care for advanced melanoma, 40–50% of melanoma cases progress while on therapies. The relationship between bacterium and carcinogenesis is well founded, such as in H. pylori in gastric cancers, and Fusobacterium in colorectal cancers. This interplay between dysbiosis and carcinogenesis questions whether changes in the microbiome could affect treatment. Thus, FMT may find utility in modifying the efficacy of anti-PD-1. This review aims to examine the use of FMT in treatment-resistant melanoma. A literature search was performed using the keywords “fecal microbiota transplant” and “skin cancer”. Studies were reviewed for …

Pediatric Parosteal Osteosarcoma Of The Distal Radius Causing Extensive Erosive Mass Effect Of The Adjacent Ulna: A Case Report, Andrea Perloff, Sehoon Park, Robert Panganiban, John Devries

Department of Orthopaedics Faculty Research

Introduction: Parosteal osteosarcomas are low-grade bony malignancies that are treated primarily with surgical resection and reconstruction. This report discusses a unique case of a pediatric patient who presented with a parosteal osteosarcoma of the distal radius causing extensive erosive mass effect and growth disturbance of the adjacent ulna. Likely due to their slow-growing nonaggressive nature, parosteal osteosarcomas have not been previously described to abut adjacent bony structures through direct contact. The patient presented in a significantly delayed manner due to social circumstances, inadvertently revealing this novel behavior. This report reviews this rare case and describes the current understanding of this …

Mob Family Proteins And Tricornered Kinase Are Required To Form Dorsal Appendages Of The Drosophila Eggshell, Keala H. D. Watson

UNLV Theses, Dissertations, Professional Papers, and Capstones

To understand how complex tissues are formed, we need to understand how cells communicate with other cells to build structures. In Drosophila melanogaster, epithelial cells form tubular structures to shape the dorsal appendages on the eggshells. One signaling pathway needed for this event is the Hippo-like kinase intracellular pathway. I focus on two key components in this pathway: the NDR kinase Tricornered (Trc) and Mob proteins. In eukaryotes, Mob proteins are known to interact with NDR kinases to promote their activation. The Hippo- like pathway involves the NDR kinase Trc; however, the specific Mob protein(s) that interact with Trc …

Characterization Of The Thermophilic Xlanase Fsa272 From Candidatus Fervidibacter Sacchari Belonging To Glycoside Hydrolase Family Gh10, Nicole Torosian, Jonathan K. Covington, Allison M. Cook, Nancy O. Nou, Jan-Fang Cheng

Undergraduate Research Symposium Posters

Candidatus Fervidibacter sacchari is a novel, facultatively anaerobic, hyperthermophilic bacterium found in terrestrial geothermal springs globally. Its genome encodes 115 putative glycoside hydrolase enzymes that are predicted to hydrolyze glycosidic bonds between carbohydrates. Fsa272, a member of the glycoside hydrolase family 10, was synthesized and cloned into Escherichia coli strain T7 Express. The transformed E. coli was grown with LB broth and ampicillin at 37°C. Fsa272 expression was induced with isopropylthio-beta-galactoside (IPTG), and the lysate was heat purified for 15 minutes at 80° C. The 3,5-dinitrosalicylic acid assay identified xylanase activity with a pH range of 4.5 to 10.5 (pHopt …

Localization Of Ipmk, Braden Sio

Undergraduate Research Symposium Posters

Inositol Polyphosphate Multikinase (IPMK) is a relatively new gene found in mammalian cells. To determine where I should further conduct our research, I localized this gene within HEK-293 cells. For this study, HEK-293 cells are used. These cells are easily cultured and transfected, both of which are needed for this study. IPMK tagged with Green Fluorescent Protein is transfected into our test group. The control group is transfected with only Green Fluorescent Protein. After transfecting the HEK-293 cells, the cells are put under a fluorescent microscope under the GFP setting. In the HEK-293 cells that were transfected with IPMK GFP, …

Data Supporting A Saturation Mutagenesis Assay For Tat-Driven Transcription With The Gigaassay, Ronald Benjamin, Christopher J. Giacoletto, Zachary T. Fitzhugh, Danielle Eames, Lindsay Buczek, Xiaogang Wu, Jacklyn Newsome, Mira V. Han, Tony Pearson, Zhi Wei, Atoshi Banerjee, Lancer Brown, Liz J. Valente, Shirley Shen, Hong-Wen Deng, Martin R. Schiller

Life Sciences Faculty Research

The data in this article are associated with the research paper “GigaAssay – an adaptable high-throughput saturation mutagenesis assay". The raw data are sequence reads of HIV-1 Tat cDNA amplified from cellular genomic DNA in a new single-pot saturation mutagenesis assay designated the “GigaAssay”. A bioinformatic pipeline and parameters used to analyze the data. Raw, processed, analyzed, and filtered data are reported. The data is processed to calculate the Tat-driven transcription activity for cells with each possible single amino acid substitution in Tat. This data can be reused to interpret Tat intermolecular interactions and HIV latency. This is one of …

Seqcode: A Nomenclatural Code For Prokaryotes Described From Sequence Data, Brian P. Hedlund, Maria Chuvochina, Philip Hugenholtz, Konstantinos T. Konstantinidis, Alison E. Murray, Marike Palmer, Donovan H. Parks, Alexander J. Probst, Anna-Louise Reysenbach, Luis M. Rodriguez-R, Ramon Rossello-Mora, Iain C. Sutcliffe, Stephanus N. Venter, William B. Whitman

Life Sciences Faculty Research

Most prokaryotes are not available as pure cultures and therefore ineligible for naming under the rules and recommendations of the International Code of Nomenclature of Prokaryotes (ICNP). Here we summarize the development of the SeqCode, a code of nomenclature under which genome sequences serve as nomenclatural types. This code enables valid publication of names of prokaryotes based upon isolate genome, metagenome-assembled genome or single-amplified genome sequences. Otherwise, it is similar to the ICNP with regard to the formation of names and rules of priority. It operates through the SeqCode Registry (https://seqco.de/), a registration portal through which names and …

Investigating Conserved Mechanisms For Embryonic Eye Regrowth In Xenopus Laevis, Cindy Kha

UNLV Theses, Dissertations, Professional Papers, and Capstones

Regeneration is a complex process to replace or restore tissue after damage. However, animals vary in their ability to regenerate tissue, and humans are limited in their abilities. Understanding how regeneration occurs in highly regenerative vertebrates, such as in Xenopus laevis frogs will lead to defining common regeneration mechanisms. In recent years, eye regeneration research in Xenopus laevis tadpoles provided new promising insight into the mechanisms necessary for repair of damaged eye tissue. Much progress has been made towards understanding the regenerative capability of the complex eye since Xenopus tadpoles can regenerate mature eye tissues, including: the optic nerve, retina, …

Trends In Utilization Of Human Papillomavirus Vaccines And The Incidence Of Malignant Cervical Cancer In The U.S. Population: A Secondary Analysis, Myah Luna, Soumya Upadhyay

Healthcare Administration and Policy Faculty Research

Background: Human papillomavirus (HPV) is a sexually transmitted infection, and HPV types 16 and 18 are responsible for approximately 66% of all U.S. cervical cancer cases in women. The quadrivalent HPV vaccine was licensed in mid-2006, and it was designed to target and protect against HPV types 6, 11, 16, and 18. The aim of this study is to examine the utilization rate of the HPV vaccine, and the trends and incidence rate of malignant cervical cancer across the United States. Methods: This study utilized data from Surveillance, Epidemiology, and End Results (SEER) and the National Immunization Survey's (NIS) teenage …

Impacts Of Motor Developmental Delay On The Inter-Joint Coordination Using Kinematic Synergies Of Joint Angles During Infant Crawling, Li Zhang, Chun F. Deng, Yuan Liu, Lin Chen, Naong Xiao, Sheng J. Zhai, Wen S. Hou, Yu X. Chen, Xiao Y. Wu

Electrical & Computer Engineering Faculty Research

Motor developmental delay (MDD) usually affects the inter-joint coordination for limb movement. However, the mechanism between the abnormal inter-joint coordination and MDD is still unclear, which poses a challenge for clinical diagnosis and motor rehabilitation of MDD in infant’s early life. This study aimed to explore whether the joint activities of limbs during infant crawling are represented with kinematic synergies of joint angles, and evaluate the impacts of MDD on the inter-joint coordination using those synergies. 20 typically developing infants, 16 infants at risk of developmental delay, 11 infants at high risk of developmental delay and 13 infants with confirmed …

Differential Expression Of Cellular And Exosomal Microrna Isolated From Oral Cancer Cells And Their Resistance To Chemotherapy, Brady Petersen

UNLV Theses, Dissertations, Professional Papers, and Capstones

Introduction: MicroRNAs are abundant small non-coding RNA with a variety of known functions, including transcriptional activation and inhibition. Recent evidence has suggested that microRNA expression may influence the responsiveness of some cancers to chemotherapy, including liver and lung cancers. Some evidence has now suggested that specific microRNAs, such as miR-21, miR-155, and miR-375, may influence oral cancer responsiveness to chemotherapy – although much remains to be discovered.

Objective: Based upon the lack of evidence in this area, the primary objective of this study was to evaluate microRNA expression and responsiveness among oral cancers.

Methods: Five commercially available oral cancer cell …

Identification Of Lipid Species: Potential Biomarkers For Early Detection Of Hepatocellular Carcinoma, Kit Yee Leung, Kaylee Freitas

Undergraduate Research Symposium Posters

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. HCC is associated with cirrhosis. Poor survival rate of HCC is due to diagnosis of HCC at late stage.

Race And Ethnic Group Dependent Space Radiation Cancer Risk Predictions, Francis A. Cucinotta, Premkumar B. Saganti

Health Physics and Diagnostic Sciences Faculty Research

Future space missions by national space agencies and private industry, including space tourism, will include a diverse makeup of crewmembers with extensive variability in age, sex, and race or ethnic groups. The relative risk (RR) model is used to transfer epidemiology data between populations to estimate radiation risks. In the RR model cancer risk is assumed to be proportional to background cancer rates and limited by other causes of death, which are dependent on genetic, environmental and dietary factors that are population dependent. Here we apply the NSCR-2020 model to make the first predictions of age dependent space radiation cancer …

Development Of The Seqcode: A Proposed Nomenclatural Code For Uncultivated Prokaryotes With Dna Sequences As Type, William B. Whitman, Maria Chuvochina, Brian P. Hedlund, Philip Hugenholtz, Konstantinos T. Konstantinidis, Alison E. Murray, Marike Palmer, Donovan H. Parks, Alexander J. Probst, Anna-Louise Reysenbach, Luis M. Rodriguez-R, Ramon Rossello-Mora, Iain Sutcliffe, Stephanus N. Venter

Life Sciences Faculty Research

Over the last fifteen years, genomics has become fully integrated into prokaryotic systematics. The genomes of most type strains have been sequenced, genome sequence similarity is widely used for delineation of species, and phylogenomic methods are commonly used for classification of higher taxonomic ranks. Additionally, environmental genomics has revealed a vast diversity of as-yetuncultivated taxa. In response to these developments, a new code of nomenclature, the Code of Nomenclature of Prokaryotes Described from Sequence Data (SeqCode), has been developed over the last two years to allow naming of Archaea and Bacteria using DNA sequences as the nomenclatural types. The SeqCode …

Xrcc4 And Mre11 Roles And Transcriptional Response To Repair Of Talen-Induced Double-Strand Dna Breaks, Ronald Benjamin, Atoshi Banerjee, Xiaogang Wu, Corey Geurink, Lindsay Buczek, Danielle Eames, Sara G. Trimidal, Janice M. Pluth, Martin R. Schiller

Health Physics and Diagnostic Sciences Faculty Research

Double-strand breaks (DSB) are one of the most lethal forms of DNA damage that, if left unrepaired, can lead to genomic instability, cellular transformation, and cell death. In this work, we examined how repair of transcription activator-like effector nuclease (TALEN)-induced DNA damage was altered when knocking out, or inhibiting a function of, two DNA repair proteins, XRCC4 and MRE11, respectively. We developed a fluorescent reporter assay that uses TALENs to introduce DSB and detected repair by the presence of GFP fluorescence. We observed repair of TALEN-induced breaks in the XRCC4 knockout cells treated with mirin (a pharmacological inhibitor of MRE11 …

Intracorporeal And Extracorporeal Anastomosis For Robotic-Assisted And Laparoscopic Right Colectomy: Short-Term Outcomes Of A Multi-Center Prospective Trial, Robert K. Cleary, Matthew Silviera, Tobi J. Reidy, James Mccormick, Craig S. Johnson, Patricia Sylla, Jamie Cannon, Henry Lujan, Andrew Kassir, Ron Landmann, Wolfgang Gaertner, Edward Lee, Amir Bastawrous, Ovunc Bardakcioglu, Sushil Pandey, Vikram Attaluri

Department of Surgery Faculty Research

Background: Studies to date show contrasting conclusions when comparing intracorporeal and extracorporeal anastomoses for minimally invasive right colectomy. Large multi-center prospective studies comparing perioperative outcomes between these two techniques are needed. The purpose of this study was to compare intracorporeal and extracorporeal anastomoses outcomes for robotic assisted and laparoscopic right colectomy. Methods: Multi-center, prospective, observational study of patients with malignant or benign disease scheduled for laparoscopic or robotic-assisted right colectomy. Outcomes included conversion rate, gastrointestinal recovery, and complication rates. Results: There were 280 patients: 156 in the robotic assisted and laparoscopic intracorporeal anastomosis (IA) group and 124 in the robotic …

Bone Morphogenetic Protein Signaling Requirement In Centripetal Migration In Drosophila Melanogaster Oogenesis, Sheila Mosallaei

UNLV Theses, Dissertations, Professional Papers, and Capstones

Cells interact with each other via signals to coordinate migration for proper tissue development and maintenance, immunity and tissue repair. Signals regulate migration via changes to cytoskeletal components or transcriptional regulation. Here, I used Drosophila melanogaster to investigate whether and how a transcriptional signal, Bone Morphogenetic Protein signaling, drives a cell migration, centripetal migration. The migrating cells, centripetal follicle cells, consist of two subsets of cells, “leading” and “following” centripetal follicle cells, that although adjacent to each other, migrate differently. The differences in modes of migration correlate to differential gene expression in the centripetal follicle cells. The leading centripetal follicle …

Recoding Of Stop Codons Expands The Metabolic Potential Of Two Novel Asgardarchaeota Lineages, Jiarui Sun, Paul N. Evans, Emma J. Gagen, Ben J. Woodcroft, Brain P. Hedlund, Tanja Woyke, Philip Hugenholtz, Christian Rinke

Life Sciences Faculty Research

Asgardarchaeota have been proposed as the closest living relatives to eukaryotes, and a total of 72 metagenome-assembled genomes (MAGs) representing six primary lineages in this archaeal phylum have thus far been described. These organisms are predicted to be fermentative heterotrophs contributing to carbon cycling in sediment ecosystems. Here, we double the genomic catalogue of Asgardarchaeota by obtaining 71 MAGs from a range of habitats around the globe, including the deep subsurface, brackish shallow lakes, and geothermal spring sediments. Phylogenomic inferences followed by taxonomic rank normalisation confirmed previously established Asgardarchaeota classes and revealed four additional lineages, two of which were consistently …

A Qualitative Review On Mammography Interventions Among Asian American Women, Sayeda Tazim Zaidi, Raisa Kabir, Chia-Liang Dai

Undergraduate Research Symposium Posters

Breast cancer (BC) is a leading type of malignancy and the second most common cause of cancer-related death among women around the world. The occurrence of BC has been increasing in Asian American (AA) women. Early detection through mammography is an effective means to reduce mortality associated with BC. However, there is a paucity of data and interventions regarding mammography promotion interventions among AA women. The purpose of this study was to evaluate current interventions for promoting mammography among AA women and suggest recommendations for designing successful interventions. A review for peer-reviewed journal articles in PubMed Central, MEDLINE, CINAHL, ERIC, …

Identifying Determinants Of Breast Cancer Screening Through Mammography Among Asian American Women, Sayeda Tazim Zaidi, Raisa Kabir, Chia-Liang Dai

Undergraduate Research Symposium Posters

Breast cancer is the most common malignancy affecting women around the world. In the US, approximately 1 in 8 women will develop breast cancer over the course of their lifetime. There is a growing incidence of breast cancer among Asian American (AA) women. Breast cancer screening by mammography is the most effective early detection method to reduce mortality but AA women have the lowest utilization rates. This study aims to identify the determinants affecting mammography among AA women. A literature search in PubMed Central, MEDLINE, and CINAHL was undertaken to include all peer-reviewed studies published within the past 20 years …