## CD-1 PFOA/GenX gestational exposure and latent health outcomes study
## Week 18 Necropsy Data
## Liver Enzymes
## HAC 12/01/2020 (w/ QC data)

## Load packages (needs to be done every time R is opened)
library(dplyr)
library(lme4)
library(ggplot2)
library(multcomp)
library(tidyverse)

# Set your working directory (replace this example with your file folder destination)
setwd("/Users/harliecope/Desktop/GenX:PFOA Study/GenX PFOA Mouse Study/GenXPFOA Study")

# Read in your files
x<- read_csv("~/Desktop/GenX:PFOA Study/GenX PFOA Mouse Study/QC Data Files/csv for R/QCWeek18NecropsyClinChem.csv")
x$ID <-as.factor(x$ID)
x$Dam.ID<-as.factor(x$Dam.ID)
x$Group<-as.factor(x$Group)

str(x)

w18necrop<-x


names(w18necrop)

w18necrop.summary.stats<-w18necrop%>%
  group_by(Group,Sex,Diet)%>%
  summarise(
    bw.mean=mean(BW, na.rm=T),
    bw.sd=sd(BW, na.rm=T),
    liv.mean=mean(LW, na.rm=T),
    liv.sd=sd(LW, na.rm=T),
    rel.liv.mean=mean(RelLivWt, na.rm=T),
    rel.liv.sd=sd(RelLivWt, na.rm=T))
w18necrop.summary.stats


write.csv(w18necrop.summary.stats, "Week18Necropsy Weights Summary Statistics.csv",row.names = F)


### Statistical Analyses

## Statistical analysis
## Males

w18m<-subset(w18necrop, Sex=="M")
w18f<-subset(w18necrop, Sex=="F")

w18mhf<-subset(w18m, Diet=="H")
w18mlf<-subset(w18m, Diet=="L")
w18fhf<-subset(w18f, Diet=="H")
w18flf<-subset(w18f, Diet=="L")

w18fveh<-subset(w18f, Group=="Control")
w18mveh<-subset(w18m, Group=="Control")
## Male, HFD
summary(glht(aov(ALP~Group,data=w18mhf), linfct=mcp(Group="Dunnett")))
## No differences

summary(glht(aov(ALT~Group,data=w18mhf), linfct=mcp(Group="Dunnett")))
## No differences

summary(glht(aov(AST~Group,data=w18mhf), linfct=mcp(Group="Dunnett")))
## No differences


## Male, LFD
summary(glht(aov(ALP~Group,data=w18mlf), linfct=mcp(Group="Dunnett")))
## Difference in 0.2 mg/kg GEnX p=0.026

summary(glht(aov(ALT~Group,data=w18mlf), linfct=mcp(Group="Dunnett")))
## no differences

summary(glht(aov(AST~Group,data=w18mlf), linfct=mcp(Group="Dunnett")))
## No differences


## Female, HFD
summary(glht(aov(ALP~Group,data=w18fhf), linfct=mcp(Group="Dunnett")))
## Differences in 2 mg/kg GenX (p=0.0325) and 1.0 mg/kg PFOA (p=0.0170)

summary(glht(aov(ALT~Group,data=w18fhf), linfct=mcp(Group="Dunnett")))
## Differences in 2 mg/kg GenX (p=0.0216) and 1.0 mg/kg PFOA (p=0.0425)

summary(glht(aov(AST~Group,data=w18fhf), linfct=mcp(Group="Dunnett")))
## Difference in 1.0 mg/kg PFOA compared to control, (p=0.0111)


## Female, LFD
summary(glht(aov(ALP~Group,data=w18flf), linfct=mcp(Group="Dunnett")))
## No differences

summary(glht(aov(ALT~Group,data=w18flf), linfct=mcp(Group="Dunnett")))
## No differences

summary(glht(aov(AST~Group,data=w18flf), linfct=mcp(Group="Dunnett")))
## No differences


summary(glht(aov(ALP~Diet, data=w18fveh))) ##diet effect P<0.001
summary(glht(aov(ALP~Diet, data=w18mveh))) ##no diet effect
summary(glht(aov(ALT~Diet, data=w18fveh))) ##no diet effect
summary(glht(aov(ALT~Diet, data=w18mveh))) ## diet effect p=0.0140
summary(glht(aov(AST~Diet, data=w18fveh))) ##no diet effect
summary(glht(aov(AST~Diet, data=w18mveh))) ##no diet effect