Saliva as a diagnostic fluid DENT 5302 Topics in Denta

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Saliva as a diagnostic fluid

• DENT 5302
• Topics in Dental Biochemistry
• Dr. Joel Rudney

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Why use saliva?

• Collection is relatively non-invasive - no
  needles!
• Range of potential analytes as broad as plasma
• Bacteria in saliva
• Salivary ions
• Secreted salivary proteins
• Inflammatory mediators
• Proteins, metabolites, from other parts of the
  body
• Nucleic acids (from the mouth and other regions)
• This is considered a high-priority research area
• The vision - chairside diagnostic testing

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The search for biomarkers

• A big buzzword in diagnostics today
• Any sort of analyte can be a biomarker
• The keys to the definition
• Changes in a biomarker must be associated with
  the active presence of a disease, or future risk
  of a disease
• There doesnt have to be a direct etiological
  relationship
• A marker for risk or diagnosis
• Multiple analytes can improve sensitivity/specific
  ity

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Earlier efforts - caries risk

• Bacteria in saliva as biomarkers - chairside
  tests
• Dentocult SM "Strip mutans", S. mutans in saliva
• Dentocult LB, Lactobacilli in saliva
• Dentobuff test strips, salivary buffering
  capacity (HCO3-)
• General problem - high sensitivity, but low
  specificity
• Combination testing (all three plus flow rate)
• Not much improvement
• These products not widely used in the U.S.

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Other oral diseases

• Detect candidal infection - Oricult N
• Not widely used in the U.S.
• Periodontitis
• The major focus is on biomarkers in gingival
  fluid
• Periodontists want to predict which sites will
  lose attachment
• Saliva can only provide indirect indications

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Saliva drug testing

• Main focus is drugs of abuse
• Much less invasive than observed urine testing
• Harder to fake, no oral equivalent of the
  whizzinator
• Products for roadside forensic testing
• Rapiscan , Oral-Screen, 4-5 drugs
  simultaneously
• Cannabinoids, opiates, meth, cocaine, diazepines
• Some published validation of these methods
• Largely unregulated online offers for home
  testing
• Matched by online offers of products to beat the
  tests
• No real validation of either

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Saliva and steroids

• Passively carried into saliva with water from
  plasma
• Accurately correlated with plasma levels
• Monitoring of female reproductive hormones
• Convenient for monitoring therapeutic uses
• Post-menopausal HRT
• Menstrual cycle timing
• Saliva and androgens
• Can be used to monitor testosterone levels
• HRT, but also by bodybuilders
• Steroids of athletic abuse (with the right test
  system)
• Also lots of online activity

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Saliva and HIV testing

• Extremely low levels of virus relative to other
  body fluids
• Assumed to be due to anti-viral salivary proteins
• SLIPI, MUC7
• Antibodies to HIV (protective or not?) are
  present
• The basis for HIV testing in saliva
• Saliva-based home HIV test on track for approval
• Controversial issues
• Pro - more people will get tested, seek treatment
• Con - no counseling, outside the public health
  system

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Sjogrens Syndrome proteome

• Early detection of salivary gland changes
• Previous studies of individual proteins
• Differences, but sensitivity/specificity
  inadequate
• Can this be improved by comparing proteomes?
• Recent data from NIDCR
• Compared to controls, patients show
• Incr. inflammatory proteins lysozyme,
  lactoferrin, cystatin C, ?2-microglobulin, Ig
  ?-light chain, polymeric Ig receptor (component
  of S-IgA)
• Decr. acinar proteins presumed PRPs, amylase,
  carbonic anhydrase
• Key question How early do these changes begin?

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Cancer antigens in saliva

• Breast cancer biomarkers
• Salivary c-erbB-2 protein - an oncogene product
• Elevated in breast cancer patients
• Sensitivity 87, specificity 65
• Proteomic studies in progress
• Not useful for prostate-specific antigen (PSA)

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Saliva and oral cancer

• The earlier the diagnosis, the better the
  prognosis
• Dentists are the first line of defense
• Oral cancer is hard to distinguish from
  transitory lesions
• When should you biopsy?
• Can pre-cancerous conditions be detected?
• Can salivary biomarkers provide an earlier
  warning?
• An area of active research at U of M

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Many potential biomarkers

• Elevated levels of certain bacteria in saliva
• Three very common commensal species
• Capnocytophaga gingivalis, Prevotella
  melaninogenica, and Streptococcus mitis
• Probably effect rather than cause
• Sensitivity 80, specificity 82
• Proteomes, genomes, and transcriptomes
• Dr. David Wong at UCLA School of Dentistry
• Collaborators at U of M
• Drs. Nelson Rhodus and Patrick Gaffney
• Dr. Kathy Moser (Sjogrens)

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Findings from the Wong lab

• Oral cancer biomarkers in the salivary proteome
• Elevated levels of the proinflammatory cytokine
  IL8
• A general marker of inflammation
• Other sources of oral inflammation - confounders?
• Additional protein biomarkers may be needed
• Oral cancer biomarkers in the salivary
  transcriptome
• Wong lab discovered that saliva contains intact
  mRNA
• Marker of changes in gene expression due to
  cancer?
• Seven genes upregulated in patients relative to
  controls
• Includes two proinflammatory cytokines, IL8 and
  IL1

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Chairside testing

• Wong lab is collaborating with engineers
• Objective lab on a chip
• Miniaturized multi-analyte detection and
  quantification
• Intended for use in a dental office
• Results at chairside
• Application Identify patients to watch or biopsy
• Detect oral cancer while it is still treatable
• The same concept can be applied to other diseases
• Oral and systemic diseases both
• People see dentists more often than physicians
• The hunt for biomarkers is ongoing

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Format of the saliva midterm

• Short-answer questions
• How Dr. Rudney defines a short answer
• When I say short, I mean short
• Some of them can be answered with a single word.
• Respond with words, phrases, or diagrams, as
  seems appropriate
• Dont mess around with complete sentences or
  essays.
• Use only the space provided for each question.
• In the case of questions worth more than one
  point
• The number of points given will be based on the
  number of correct answers you provide for that
  question
• When I ask for examples, it means that the number
  of potentially correct answers is greater than
  the number of examples that Im asking for.
• There are no trick questions (at least, not
  intentionally)

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A sample one-point question

• Q. Identify the component of the water secretion
  system that requires energy to function. (1 pt.)

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A sample one-point question

• Q. Identify the component of the water secretion
  system that requires energy to function. (1 pt.)
• A. Na-K-ATPase
• You get one point

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A sample multi-point question

• Q. Give three examples of proteins found in
  pellicle (3 pts.)

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A sample multi-point question

• Q. Give three examples of proteins found in
  pellicle (3 pts.)
• A. Statherin, acidic proline-rich proteins
  (aPRP), histatins
• You get three points

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A sample multi-point question

• Q. Give three examples of proteins found in
  pellicle (3 pts.)
• Acceptable answers
• Statherin, acidic proline-rich proteins, amylase,
  histatins, cystatins, MUC7 mucin, lysozyme,
  albumin, carbonic anhydrase
• Use any three, and you get three points

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A sample multi-point question

• Q. Give three examples of proteins found in
  pellicle (3 pts.)
• Acceptable answers
• Statherin, acidic proline-rich proteins, amylase,
  histatins, cystatins, MUC7 mucin, lysozyme,
  albumin, carbonic anhydrase
• Your answer
• Statherin, aPRP, basic PRP
• You get two points