---start path.lec.04.29.97----

dr weber again! woo hoo! (or something)

the exam- everyone did well on it, avg around 90 or so. (94)
final is worth 60% though, so don't "slough off" - keep on working... 
etc.
Dr chacko and weber have a major concern - they spent 45 min transferring 
grades to a master sheet...and the handwriting on the exams SUCKS when it 
comes to the names. ten had to be done by process of elimination, so from 
now on PRINT your names, LAST name first, it must be CLEARLY LEGIBLE.

ok.
slide: cute little bulldog type puppy :)

moving on with the neoplasia stuff...
reminder...we're talking about development of neoplasm...uncontrolled 
growth of cells and loss of regulation that normally prevents excessive, 
continuous dividing. remember some main points re: what normally turns 
cell on...
growth factors, GF receptors, transducers, all going into common pathway 
leading to phosphorylation of proteins, and the phosphorylation 
ultimately gets into nucleus, and you start having cells enter cell 
cycle. one particular type of gene product Myc only functions in nucleus, 
is simply a DNA regulatory protein,a nd by binding to DNA puts cell into 
mitotic cycle.

how does each oncogene affect the cell?

GROWTH FACTOR GENE: regulates production of a protein product. sis 
(simian sarcoma) codes for something very like PDGF....if sis is changed 
into c-onc from p-onc, may  start to make INCREASED GROWTH FACTOR 
PRODUCTION. cell will start seeing more factor on the receptors, and so 
more turnover can occur. if you consider osteosarcoma or something where 
growth factor is involved, there could be autocrine stimulation of tumor 
by cell product made by tumor cells themselves....bottom line, you just 
have EXTRA GF causing overstimulation of local cells.

INCREASED GROWTH FACTOR RECEPTORS on cell surface can also occur. erb-b 
or c-neu...causes increased growth factor receptors to be expressed, 
therefore, cell is exquisitely sensitive to the normal GF. can respond to 
very low concentrations where before it might not have responded, so cell 
can be activated abnormally.

TRANSDUCER MUTATIONS: very common, found in almost all human tumors in 
many tissues. a mutation in the ras signal transducer involves this 
mutant signal transducer protein constantly signaling the cell to enter 
cell cycle.  everything else is normal

fig 7-17 in handout...
note that normal gene product of ras is p21. the normal p21 gene product 
normally binds GDP, the cell is signalled by a surface receptor (eg GF 
receptor, NT, whatever) and GDP is converted to GTP, activating ras, 
causing hydrolysis of GTP, ATP, cAMP, cAMP kinase, etc etc....normal 
process. BUT if ras gene is mutated, it forms a protein that primarily 
binds the GTP not the GDP. ADDITIONALLY, the normal protein has GTPase 
activity, so once GTP is bound, it is hydrolyzed and drives the p21 back 
to the GDP form, and cell stops dividing. in mutant form, there is a lack 
of GTPase, so activated ras protein stays around, can't go back to GDP 
binding form, so cell constantly activated.

but remember takes more than one mutation to transform cell into 
malignant state.

mutant transcription factors:
mutant myc - enters and binds DNA in nucleus, turning cell over 
continuously
myc is a mutation very frequently found in many tumors, like ras 
mutation.

so far, we've discussed only p-oncs and their mutations into c-oncs, 
which generally increase cell turnover and keep cell in cell cycle. but 
we also need to lose the effects of the suppressor genes. the suppressor 
genes are intended to inhibit cells from entering cell cycle. the rb 
retinoblastoma gene has been looked at a lot. we come to a situation...we 
are ending up with a tumor in the eye which is based on transformation of 
retinoblasts. how does this tumor originate? two forms. familial, and 
sporadic.

familial rb == 40%, sporadic == 60%.

if you have a male who has one mutant RB gene, and one normal gene, then 
all of his children will have one of the mutant genes, (why? not sure. 
but thats what he said) but at that point, there is no evidence of any 
tumor or malfunction. you have to also have another mutation...a somatic 
mutation causing homozygosity - you need loss of normal rb gene in the 
offspring who already carry one mutant allele. this is the "two hit" 
phenomenon. loss of single allele is not sufficient.


in sporadic tumors, children are all normal, have no inherited mutant 
alleles. you need TWO mutations...a sequence of mutations must occur, to 
KO both suppressor genes..

how does this RB gene function, and why does cell enter cell cycle when 
RB is lost? if you consider normal cell cycle...normally, a retinoblast 
just sits there, doesn't normally divide, is in stable category.  when RB 
is not phosphorylated (eg, at end of m phase) cell stays in G1 resting 
state, or G0 state. but if RB is phosphorylated - eg pRB-P, which occurs 
at end of G1, this allows cell to enter cell cycle, S phase. so the LOSS 
Of RB would somehow then allow cell to enter cell cycle.

some oncogenic DNA viruses inactivate RB by binding to it in 
unphosphorylated state, which knocks that gene out, so it can't do its 
job. cell would then be continuously driven through cycle, b/c RB can't 
be in nonphosphorylated state. must have RB in non-phosphorylated state 
to keep cell in G1/G0.

another suppressor gene functions similarly. that is p53, a commonly 
mutated gene in many neoplasms. several possibilities. if mutagenic 
agents affect a normal cell, you know that cells can repair DNA damage to 
some extent w/enzymes that splice it and fix it, etc, and w/o those 
enzymes, we'd have more problems than we do today. but if you have these 
mutations in normal cell, sometimes p53 is increased, and cells are 
arrested in G1, DNA can then be repaired and return to normal, OR, if 
not, can go into apoptosis and can die. this happens as long as p53 is 
increased, and there is no mutation in p53. BUT if there is a mutation in 
p53, resulting in loss of suppressor gene product, you have no rise in 
p53 after mutagens attack cell. then, cell can't repair damage anymore, 
but rather, the mutated cells start to proliferate, and become 
transformed cells. you've lost the p53 suppressor functions which 
normally keep the cell from cycling.

so when you have a neoplasm developing, it is a SEQUENCE OF CHANGES In 
activation of p-onc-->c-onc and in loss of suppressor gene function.

colon carcinoma in humans...you start with normal epithelium, then you 
have loss or mutation of APC locus (a suppressor gene) and you get 
hyperproliferative epi, then loss of DNA methylation and you get early 
adenoma, then mutation of ras and bigger adenoma, then loss of DCC and 
late adenoma forms, then loss of p53 and you have adenocarcinoma. so you 
see there are many mutations occuring...just realize there is sequential 
loss of at least 3 suppressor genes, and activation of one p-onc becoming 
a c-onc (ras). so this is a good example of how neoplasia develops.
furthermore, the precise sequence of events isn't actually as important. 
the NUMBEr of mutations is more important. you get a cumulative effect.

one other thing re: suppressor genes. they don't only function via 
phosphorylation. eg, DCC gene produces something on cell surface, an 
adhesion molecule that is lost. so, loss of suppressor gene could cause 
loss of adhesion molecule, and that would cause poor cell-cell contact, 
cells would be more migratory, would get the ability to infiltrate other 
local tissues b/c less adhesive to it's own tissue type...so not only 
phosphorylation is a big deal. many processes are affected.

end of p-onc and suppressor discussion.

slide: black wartlike thing, hairless. to me, this looks pretty well 
demarcated. it's pretty round, shiny, protruding above skin surface. at 
bottom of slide, possibly some irregularity of margin. NOTE: any time you 
see a black tumor mass, in man or animal, you always worry, even if it 
looks benign, because it is most likely going to be a malignant melanoma. 
even though melanoma CAN be benign, they tend to METASTASIZE very very 
early, and should arouse tremendous concern.

histo: how can you positively ID malignancy vs benign quality of tumor? 
here we see a LOT of pigment production. cells making the pigment are 
very very dark, we can't really see nuclei or anything. cells NOT making 
pigment on this slide do look reasonably uniform. no mitotic figures. 
some oval, some round, but reasonably well differentiated, b/c making 
lots of pigment, so has normal function.  if cells are making pigment, 
can't be undifferentiated/anaplastic, b/c then wouldn't be making 
pigment. so, i'm not sure what he's calling this one..

slide: mucocutaneous jct lip of dog. multinodular subq projections of 
black masses, very irregular border, not well defined at all. darker 
discolorations in some areas beneath the furn. partly hairless. partly 
ulcerated.

slide: sag section - local mets to retropharyngeal area from above tumor. 
clearly we see invasion of normal tissue by this abnormal black tissue, 
extendign fingerlike projections with ambiguous borders. 

slide: lung with large mass in one lobe, and multiple pinpoint black sl 
protruding nodules in other areas. another nodule is partly black, partly 
grey, with some vascularity. why is this part unpigmented? well, perhaps 
it is more anaplastic? most likely, some tumor cells in those areas are 
less differentiated, more aggressive, anaplastic, and pleomorphic. they 
still make a little pigment, but WAY less. this great reduction in normal 
function indicates reduced differentiation.

slide: kidney of this dog. multiple cm sized black, well defined nodules 
in kidney cortex. now, the borders are well defined which is 
characteristic of benign tumor, but these are not benign tumors. you 
can't have multiple black benign tumors of the kidney. melanoblasts don't 
BELONG in the kidney. we've just had tumor cells lodging here in the 
vasculature.

also in brain, spinal cord (poorly pigmented)

histo: sporadic pigment producing cells. not as much as should be there. 
nuclear:cytoplasmic ratio is relatively normal, but mitotic figures (very 
condensed nuclei) are present. - check poster in lab. not always do 
malignant tumors show EVERY possible aspect of pleomorphism. not always 
are there huge differences in size/shape of cells and nuclei..but we will 
see SOME of the characteristics. in this case, we have lack of 
differentiation (anaplasia) and we have a high mitotic index - huge 
increase in mitotic figures just doesn't belong in benign tumor, so this 
MUST be malignant neoplasm.

slide: big pink blob. he says it is a rib but looks like a big ugli fruit 
to me.
he says it is a "very large, probably 15 cm x 8 cm slightly multinodular 
red mass" that appears well bordered and in fact appears encapsulated. so 
the question is, what kind of tumor might this be? no other tumor in 
animal. this one was encapsulated. so personally, hey, it sounds benign 
to me, but what the heck do i know? he says it is very firm, almost hard. 
could be osteoma, chondroma, fibroma, etc. it's at the costochondral 
junction. it is in fact a chondroma.

slide: chondrosarcoma of stifle joint. the tumor mass is not clearly 
outlined, there is local destruction of tissue...tumor has totally 
occupied joint space, eroding and destroying the articular surface of 
bone. has extensively infiltrated the area, there are cartilaginous 
masses in the whole joint capsule and kind of smushing into the bones.

slide: weird ass looking lungs. all lobes are affected by multifocal red 
to black millimeter to centimeter raised nodules. looks to me like 
hemangiosarcoma. i guess could also be malignant melanoma. he says this 
is prime example of "shower of tumor emboli reaching lung and growing 
into multiple masses". so tumor cells come in via blood vessels and 
settle in and grow. probably all showed up at same time, since similar in 
size. if only ONE lobe is involved, could be primary tumor, but this 
really looks like metastatic dz. so where could this have originated? I'd 
guess maybe spleen. why? i dunno.. i guess actually, I would say maybe 
this is a right atrial mass - a right atrial hemangiosarcoma, saieth 
doctor gorman :)
ok, he is showing us a spleen, which he says is an excellent place for 
hemangiosarcoma to start, or for it to metastasize to. this spleen has 
multiple hemorrhagic masses. now, he's showing us a heart. some breeds 
like GSD get right atrial (aha!) primary hemangiosarcomas, which btw can 
rupture - if that happens suddenly, that is very bad news for the dog, as 
pericardial sac fills w/blood and immediately compresses heart and dog 
would die w/o immediate intervention (and probably with it). but before 
actual rupture, can cause slight shedding of tumor cells, and get 
attachment of tumor cells onto epicardial surface b/c they flaked 
off...this is metastasis by SEEDING. when tumor is in a space, you can 
get implantation by seeding...this is w/o going through blood 
vasculature, but rather by moving through a local space and implanting 
nearby.

----break-----

more slides...

he's trying to cover some more practical stuff...

ear of white cat. distal pinna has poorly defined, granular, black, 
hairless mass with considerable tissue destruction at proximal border of 
mass. what is this dark mass? now, white cats do not MAKE melanin. but 
they DO get squamous cell carcinomas....UV light (sun) frequently affects 
skin at tip of ears, nose, in nonpigmented areas (also in hereford cows 
which are outside a lot). so this is a squamous cell carcinoma. could 
this be the result of trauma? highly doubtful. doesn't look like 
exuberant granulation tissue with a lot of hemorrhage to me, but dr weber 
says it COULD be. you have to consider all tissue that could be 
invovled...adnexal structures, cartilage, fibroblasts, etc. most likely 
to be involved is labile tissue like EPITHELIUM. this is scc. it's black 
due to tissue destruction, oozing of blood, etc. the blackness is crusted 
blood on the surface of this growing tumor mass. so don't think black is 
ALWAYS pigment in the tumor. could be blood.

slide: white cat w/nonhealing ulceration on tip of nose. there is 
considerable ulceration and hemorrhage there. owner brings it in for this 
nonhealing lesion. it won't heal, because tumors do not respond to 
topical treatments such as you'd use on a skin wound. this is an invasive 
neoplasm, an scc as above. it's not a very good place to operate, either 
- youd end up with a noseless cat, which most people do not want to end 
up with.

wrt SCC...where is it most likely to metastasize to? well, locally 
draining LNs are most crucial place to look for carcinoma mets. most 
carcinomas, first place to look for mets is regionally draining LNs b/c 
carcinomas tend to metastasize via the lymphatics primarily. now, 
SARCOMAS eg tumors of CT like osteosarcom and chondrosarcoma, tend to met 
via hematogenous route, invade venous system and go that way. but can go 
other way. this is a generalization.

histo: scc - surface epi with CT beneath it. islands of epi cells, 
marginally differentiating and producing some keratin "pearls" within the 
tissue. this is automatically a malignancy because if it were benign 
tumor of epithelium, would be a papilloma growing out of skin surface. 
but this epithelium has invaded the local tissue. is reasonably well 
differentiated, but it has infiltrated and invaded and locally destroyed 
the connective tissue, even though it's well differentiated with few 
mitotic figures

tumors of glands are hard to figure out sometimes. 
slide: thyroid gland, intact
slide: thyroid gland, sectioned.

intact gland is enlarged, well defined, dark tissue mass with clear 
border. some discoloration on sectioned gland. on section of intact 
gland, we see dark almost black, bulging tissue mass which has occupied 
and replaced normal gland space, but has stayed within confines of gland. 
what neoplasms occur in thyroid gland? could be thyroid epi cells, 
possibly blood vessels, fibroblasts...this one is a THYROID 
ADENOMA...tumor of thyroid tissue, of the acinar cells. why is it so 
dark? endocrine tumors, tumors of gland tissue, frequently quite 
vascular, because endocrine structures are supposed to be vascular. this 
mass is so dark b/c there are a lot of blood vessels present. has nothing 
to do w/pigment or anything. 

histo: these tumors are identified by what they are doing. there is no 
colloid - minimal colloid seen, but the cells are in acinar structures 
and look pretty normal, so it's an adenoma. BUT...you might get into 
situation where you suddenly find similar cells in say, a lung. so these 
"benign" thyroid adenomas can metastasize very early, and then you have 
to call it malignant, because it's invading the lung. 

histo: lung alveoli with discrete "thyroid adenoma" nodule present 
adjecent to normal alveoli.

so, you have to call this an adenocarcinoma of the thyroid.


off to lab....
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