The present study aimed to predict whether Holstein Friesian cows are at risk of ketosis during the transition period by analyzing hematological and serum biochemical parameters on the calving date. To the best of our knowledge, this study is the first to identify the associations of hematological and serum biochemical parameters on calving date with ketosis during the postpartum transition period. We observed that elevated MCV, MCH, NEFA and TB were positively associated with ketosis and its severity. The cows with greater RDW, WBC count, monocyte count, eosinophil count, ALB, ALT, LDH, and amylase were at lower risk of ketosis and its severity. MCV, MCHC, lymphocyte count, and eosinophil count were significantly different between subclinical and clinical ketosis.
Red blood cell indices are associated with body dysfunction. However, only a few previous studies investigated the association between ketosis and red blood cell indices in cows, and they showed no association between ketosis and MCV, MCH, MCHC, and RDW 13,22; the finding is different from that in the current study. As shown in Table 1, the incidence of ketosis varied during the transition period. The discrepancy between our findings and the previous findings may stem from differences in study design, such as monitoring frequency. The liver is an organ closely related to ketosis. Humans with liver disease and deficiencies in vitamin B9 or B12 showed elevated MCV and MCH 23,24. Dairy cows with elevated MCV and MCH have a weak liver, which may lead to ketosis. RDW is known to be elevated in patients with liver disease. Interestingly, the RDW findings in this study are inconsistent with those in previous studies on liver diseases; the reason is unclear, but a previous study showed a similar association between RDW and the severity of ketosis in sheep, although it was not significant 25. Our findings suggest that unlike cows with other liver diseases, those with severer ketosis have lower RDW values.
Accumulating studies indicate that ketosis is associated with decreased WBC count, and ketosis increases the risk of diseases. Dairy cows with ketosis show decreased WBC count 14,26. Human adults with ketogenic diets show a significant decrease in WBC count 27. Ketosis increases the risk of metritis and mastitis 28. Consistently, the present study found lower WBC counts and severer ketosis during the transition period. The decrease in WBC count in cows with ketosis might contribute to the susceptibility to infectious diseases. NK cows had extremely significantly more monocytes than CK cows and significantly more monocytes than SCK cows. Both cows and humans with hyperketonemia have lower monocyte counts 29,30. Humans in the low-energy state have a reduced number of circulating monocytes migrated from bone marrow, by signaling from hepatocyte 31. The liver might already play a role in recruiting a smaller population of monocytes from the bone marrow before ketosis in the ketosis groups. In addition, NK cows showed very significantly more eosinophils than SCK cows, while SCK cows had significantly more eosinophils than CK cows. Human with ketosis also showed a lower population of eosinophil 32,33. It is speculated that eosinophil count is lowered by oxidative stress, elevated apoptosis, and the inhibition of cell proliferation 32.
As for serum biochemistry, most of the parameters associated with ketosis (NEFA, ALB, TB, ALT, and amylase) are highly related to hepatic dysfunction. A negative energy balance in animals promotes lipolysis, which mobilizes NEFA from the adipose tissue to most body tissues through the blood stream. The liver is the main organ to process NEFA into 3 forms: ATP via complete oxidation, ketone bodies, or very low-density lipoproteins via incomplete oxidation 4,34. The liver of ketosis cows may not be able to remove NEFA compared to non-ketosis cows around the parturition before ketosis onset. In addition, ALB and TB are commonly used to indicate damage to the liver, and our results on ALB and TB correspond with previous findings. CK, SCK, and NK had lower serum ALB levels in order, which indicate from hepatic dysfunction. 21,35. TB increases in ketosis, indicating liver damage 18,36, and a higher level in TB in parturition suggests hepatic dysfunction. Notably, the ALT results in the current study are different from those in previous studies, which demonstrated that ALT is insensitive in ruminants and cows with ALT levels 21,37,38. The difference might be due to the study design. Hepatic apoptosis is elevated in dairy cows with ketosis 39. Vaden et al. showed that low levels of ALT indicate hepatic atrophy, while high levels of ALT indicate hepatocellular injury or leakage 40. CK cows had the lowest level of ALT while NK cows had the highest on the parturition day. The findings suggest that the extent of hepatic atrophy in Holstein cattle is linked to ketosis.
To the best of our knowledge, no study has reported an association between serum amylase and ketosis in cattle. However, low concentrations of serum amylase are associated with the degree of liver dysfunction, such as non-alcoholic fatty liver, metabolic syndrome, and insulin resistance 41,42. In human medicine, a previous study revealed that low serum amylase levels were inversely related to serum ketone body 43. Insulin resistance is also related to ketosis in dairy cows 44. Our findings suggest that ketosis may be associated with low serum amylase concentration.
LDH is known to contribute glycolysis, and the elevated levels of LDH are a marker of organ damage, including muscles and liver damage 45. LDH was expected to be higher in cows with ketosis because other biomarkers indicated obvious damage to the liver. However, we found that LDH had a negative relationship with the severity of ketosis. Information on the association between LDH and ketosis is limited. Energy production might presumably be related to our unexpected findings that NK had the highest level of LDH and CK had the lowest level of LDH. Indeed, LDH plays an important role in the regulation of glycolysis 46. Lower level of LDH in ketosis might indicate a decreased ability to use glucose for energy, which might lead to fat mobilization.
The limitation of our study is that it was conducted at only one farm. However, the current study demonstrated that the parameters of the complete blood count (CBC) test and serum biochemistry on the calving date were associated with ketosis and its severity. The parameters were statistically significantly different based on the severity of ketosis. Our findings suggest that theses parameters are predictive indicators for ketosis.
In summary, the present study found that hematological and serum biochemical parameters on the calving date were associated with ketosis and its severity during the postpartum transition period. Dairy Holstein cows with clinical ketosis showed significantly the highest values of MCV, MCH, NEFA, and TB but significantly lowest values of RDW; WBC, monocyte, and eosinophil counts; and ALB, ALT, LDH, and amylase levels. Dairy Holstein cows with non-ketosis showed the opposite results. Our findings suggest that dairy Holstein cows, that are susceptible to ketosis, can be selected by hematological and serum biochemical parameters.