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• The test includes 15 questions

mRNA Vaccines

• The mRNA for a spike protein of interest is extracted
• This mRNA is then placed into liposomal vesicles
• After injection, cells create the intended protein

Issues with mRNA

• mRNA could cause immune reactions or degrade
• One solution is to change a few of the mRNA bases, creating modified mRNA
• mRNA is delivered into cells via lipid vesicles

mRNA vs Traditional Vaccines

• Traditional vaccines use live/weakened or dead pathogens
• Traditional vaccines are slow to produce antigens of interest
• Immune response is stimulated when injected into the arm
• mRNA vaccines utilize mRNA, which is easier to produce
• The body produces a protein of interest, triggering an immune response

Plants

GVA

• A chemical component discovered in dill and parsley
• It inhibits mitotic division and can be used in cancer treatment
• GVA is not cytotoxic to healthy cells

Photosynthesis

• An ideal process for plants that has not changed much over evolution
• RUBISCO, a major enzyme in photosynthesis, sometimes uses O2 instead of CO2
• This results in glycolate as a byproduct, which spreads throughout the cell
• Researchers modified genetic info to keep glycolate confined, using tobacco plants
• This significantly increased plant biomass

Photosynthesis in Mammalian Cells

• NTUs (nano thylakoid units), mini-engineered chloroplasts, were placed into mice with osteoarthritis
• The condition improved over time
• NTUs cause a significant increase in NADH and ATP, even in mammalian cells

Plants for Drugs

• Biologics are drugs made by genetic engineering or plants
• They are normally done with mammalian cells
• Elelyso, a treatment by Pfizer for Gaucher disease only uses plant cells
• More funding is now going towards plant biopharmacology

Organ Transplants

Xenotransplantation

• Taking cells or organs from one species to transplant into another
• This has been done with pig kidneys, which were genetically modified using CRISPR to lower the chances of immune rejection
• The receiver of the kidney survived for 2 months

Tissue Engineering

• A sub-discipline of regenerative medicine involving building tissues in the lab using somatic, embryonic, adult, or STEM cells
• Tissue engineering assembles functional constructs that restore, maintain, or improve damaged tissues/organs
• Tissue engineering has been used to fix internal organs to produce Paneth cells
• Paneth cells produce defensins that protect the organ from pathogens
• Missing or damaged Paneth cells characterize intestinal diseases (IBS)

Organoids for Research

• Multi-chamber organoids resemble the heart and are used for research to understand cardiovascular diseases and heart development

STEM Cells

• Mice STEM olfactory cells were placed in rats and restored smell
• STEM cells could potentially restore degenerated senses in humans

Nanomedicine

Self-Boosting Vaccines

• Vaccines providing booster doses at specified times
• Use nanoparticles is helpful in areas without sufficient medicinal resources

Nanobots

• Tiny magnet-controlled robots can reach deep into lungs, where treatments cannot
• Less invasive compared to chemo

CRISPR

• A gene-editing tool, initially used for lab work, averted a blood disease in an embryo
• Casgevy is a CRISPR treatment for sickle cell

Synthetic Biology

• Involved in creating complex, biologically inspired systems with functions outside of nature

Optogenetically Responsive Mitochondria

• Mitochondria made more light sensitive were developed in vitro
• Mitochondria with this trait produce more energy when exposed to light because of a light-sensitive photon pump
• In vitro testing showed that organisms with these modified mitochondria live longer

Lab on a Chip

• Mimics human organ function and allows disease study without harming anyone
• Can mimic heart attacks and replicate the blood-brain barrier

Inherited Disease

DNA Tests

• Not all are FDA-approved, and some offer non-diagnostic medical results, like 23 and Me

Intracellular Nuclear Injection

Biomarkers

• Biological molecules in blood or urine indicating normal or abnormal body functions, such as proteins, CTCs, and cell-free DNA (cfDNA)

Biopsies

Liquid Biopsies

• Use urine or blood (mainly blood), taking a few days for results.
• Blood contains cfDNA, which is good for diagnosing cancer and readily available

Skin Biopsies

• Take six weeks for results, cannot be done at a regular medical office, and cannot detect cfDNA

Galleri Test

• Analyzes cfDNA from 50 types of cancer, is very accurate, costs over $1000, and returns results in 2 weeks

Theranos

• A biomarker company that claimed to detect blood disorders, but faked their data and was known as a scandal

CTCs (Circulating Tumor Cells)

• Responsible for metastasis
• Cynvenio uses antibodies to count CTCs
• Vortex, an improvement, separates CTCs by properties, like how WBCs and RBCs travel quicker, inspired by rivers

Cancer Stem Cells

• Multiply indefinitely, resistant to chemotherapy, responsible for relapse, and very low in number

CTC Exosomes

• A better way of measuring cancer progression than CTCs, with devices being made for counting them

ITEARS

Cancer Therapies

Tumor Microenvironment

• If the signaling is properly maintained, it allows for idealization, which prevents tumors from "waking up" and growing
• Dormant CTCs require years to metastasize and cannot reactivate in a tumor microenvironment with type III collagen

Using the Immune System

• Uses antibodies, modified T-cells, and immunomodulators (allowing cancer cells to signal apoptosis)

Biotechnology

• A discipline of developing medicinal solutions that benefits human health and welfare

Personalized Medicine

• Treating patients based on their own genetics, such as mapping for $1000, to determine lifestyle adjustments
• Testing via Genesight, Myriad genetics, and GenomicHealth
• Covered by GINA (Genetic Information Nondiscrimination Act), which prevents medical DNA tests from affecting personalized medical test results affecting someone’s employment or insurance.

Orphan/Rare Diseases

SMA (Spinal Muscular Atrophy)

• The leading genetic cause of infantile death
• Impacts the whole family and affects 1/6000 babies worldwide
• Caused by faulty genes that impact muscle nerve connections, and treatments are very expensive

FOP (Fibrodysplasia Ossificans Progressiva)

• Turns cartilage into bane, has a known genetic cause, but has no treatments

Bioethics

hESCs (Human Embryonic Stem Cells)

• Derived from human blastocysts, requiring their destruction
• Used in STEM cell therapies (allowed in US clinical trials but cannot receive federal funding if used/destroyed)

Synthetic Embryos

• In vitro embryos used to bypass hESC regulation
• Behave like real embryos, can grow a fetus when implanted in monkeys
• It is still debated whether they are considered life

Cloning

• Dolly the sheep had health issues(none of her clones did), federally permitted but states can impose regulations

Aging

Multi-factorial

• Includes mitochondrial and epigenetic factors, Telomeres
• Sirtuin genes (Longevity genes); overexpression increased fruit fly lifespans 50%
  • SIRT 6, when overexpressed in mice, increased lifespans 30%, better usage of energy from fatty acids, and fewer age-dependent cancers/disorders
• Resveratrol stimulates human sirtuin genes but is not for clinical use

Stem Cell Theory of Aging

• Younger people's stem cells function better and are more plentiful Longeveron uses mesenchymal stem cells (MSC) in potential aging therapy research
• US military explores anti aging pill with NAD+ enhancer for more mitochondrial energy, working similarly to sirtuin genes

Accelerated/Slowed Aging

• Epigenetic factors play a role, with chemicals possibly speeding or slowing aging via epigenetic influence

Random Toxins

Botulism

• Used in Botox, acts as a muscarinic ACTH inhibitor

RNAi (RNA Interference)

• Causes gene silencing by stopping mRNA from translating certain diseases
• Used for various treatments
  • Cancer, macular degeneration, Huntington’s disease

Ricin

• From castor beans, shows potential in cancer as a conjugate antibody
• Used in in vitro toxicology for discovery

Tissue Engineering & Regenerative Medicine

Tissue Engineering

• Lab-grown tissue for body area patches

Stem Cell Therapy

• Uses stem cells to treat diseases

Gene Therapy

• Uses gene transfection to treat diseases

4 Types of Stem Cells

• Adult stem cells
  • AdMSCs from fat tissue are most popular for trials, but less ethical

hESCs vs Adult Stem Cells

• hESCs are easy to produce and more pluripotent
• Adult stem cells can be sourced from hair, fat, and teeth
• Adult stem cells are analogous (can use one’s own stem cells), but can't with hESCs

Conclusion

• Cell biology blends religion, science, government, bioethics, politics, drugs, business, and regulation

Intro to Microscopy

• Microscope’s are the foundation of cell bio and was invented by Hooke in 1650s
• Leeuwenhoek is the father of microbiology (1670s)
• Schleiden and Schwan developed modern cell theory (1830s)

Brain

• Reticular theory proposes neurons aren't individual cells
• Neuronal theory states neurons are cells

Resolving Power

• Refers to smallest distance that a microscope can discern
• Light microscope : Plant (100 µm), animal cells (10μm), mitochondria (1µm)
• Electron microscope: Viruses/ribosomes (100nm), proteins (10nm), molecules (1nm)

Resolution

• Denotes the capacity to distinguish between 2 objects Light microscope: 0.2 µm
• Electron microscope: 2.4 Å
• Abbe's equation helps find theoretical limit
• Distance = 0.61*wavelength/n sinø, standard distance = ½ wavelength

How to Get Better Contrast

Contrast

• Differentiates between sample/background for detail:
• Cells mostly water with little contrast
• Organic compounds absorb light, heat, and shift around

Types of Microscopy

Bright Field Microscope

• Used for dead cell pathologist by fixating with chemicals such as formaldehyde to crosslink proteins
• Kills (fixates" the cells then dehydrates using xylene

Faster option vs Regular Microscopy

• Cryosection: quick sample freezing(no paraffin), used for Mohs surgery(skin cancer removal)

Phase microscopy

• Uses unfixed cells
• Uses light interference for better contrast and is used mainly by cell culture biologists

Differential Interference Contrast (DIC) Microscopes

• Shows living cells with 3D image and is used for single cell electrophysiology
  • Looks at neuronal drug response

Dark Field Microscope

• Highlights brighter images on a dark background and is mainly used by biologists

Confocal Microscopy

• Computer used to create 3D models of fluoresced cells

Advantages

• Less stray light
• Point to point gathering allows precise imaging

Problems

• Fluorochromes can photo bleach (this is normal)

Fluorescence applications

• Dyes must stay alive
• Measure and analyze changes in cell behavior
  • Dye can show intercellular function without being the cell

TIRF Microscopy

• Intracell inject via lucifer yellow
• Dye will travel throughout cell using ions
• Mainly for neurologists to see connectivity junctions
• Dye has to be able to cross a line without killing a cell

Immunocytochemistry

• Uses antibodies to distinguish protein in a cell
• One branch connects to antigen and one brand connects to fluorescent dye for antigen

Making antibodies

• Polyclonal antibodies made by many cells in body, injected in an animal and blood is collected
• The animal eventually stops producing

ELISA

• Enzyme linked immunoassay is used to quantify protein
• Protein tested is isolated: enzyme’s react
• Is placed and spectrometer and adjusted

Cell Death

Necrosis

• Pathological cell death, due to external causes causes the cell to explode

Apoptosis

• Genetic cell death, due to internal causes that causes the cell to implode

Fluorescent Proteins

• Acts like dyes but synthesized in vitro
• Green Fluorescent Protein (GFP) introduced and developed to indicate what is happening in the cell

Biosensors

• Protein that reveals change in cell behavior

Autoradiography

• Radioactive probe to see cell processes

FISH (Fluorescence in Situ Hybridization)

• Degradation of DNA and RNA

Cell Separation

• Calcium is used for cells to adhere for homogenous cells
• Separating clump of cells into several singular cells is Cell dissociation

FACS (Fluorescence-Activated Cell Sorting)

• Utilizes a range of antibodies for different cell types to test the protein
• Cells placed in ficoll as a distinguishable gradient

ELISA (enzyme-linked immunosorbent assay)

• Isolating insulin
• Veridex: tests if a cancer spreads

Cell Fractioning

• Breaks up cells of interest (cell lysis) to look at different sections

Cultures

• Brain tested that neuronal was true

Stem cells

• Oncongennically transformed (mutated) can grow forever