Consumption of the cell-free or heat-treated fractions of a pitched kefir confers some but not all positive impacts of the corresponding whole kefir

Abstract

Introduction: Kefir consumption can have many metabolic health benefits, including, in the case of specific kefirs, improvements in plasma and liver lipid profiles. Our group has previously shown that these health benefits are dependent on the microbial composition of the kefir fermentation, and that a pitched kefir (PK1) containing specific traditional microbes can recapitulate the health benefits of a traditional kefir. In this study we investigated how different preparations of kefir impact cholesterol and lipid metabolism and circulating markers of cardiovascular disease risk and determine if freeze-drying impacts health benefits relative to past studies.

Materials and methods: Eight-week-old male and female C57Bl/6 mice were fed a high fat diet (40% kcal from fat) supplemented with one of 3 freeze-dried kefir preparations (whole kefir, cell-free kefir, or heat-treated kefir) for 8 weeks prior to analysis of plasma and liver lipid profiles, circulating cardiovascular disease (CVD) biomarkers, cecal microbiome composition, and cecal short-chain fatty acid levels. These groups of mice were compared to others that were fed a control low-fat diet, control high fat diet or high fat diet supplemented with milk, respectively.

Results: All kefir preparations lowered plasma cholesterol in both male and female mice, while only whole kefir lowered liver cholesterol and triglycerides. Plasma vascular cell adhesion molecule 1 (VCAM-1) was lowered by both whole kefir and heat-treated kefir in male mice but not females, while c-reactive protein (CRP) was unchanged across all high fat diet fed groups in males and females.

Conclusion: These results indicate that some of the metabolic benefits of consumption of this kefir do not require whole kefir while also indicating that there are multiple compounds or components responsible for the different benefits observed.

Keywords: cardiovascular disease; cholesterol; kefir; metabolic health; non-alcoholic fatty liver disease.

Figure 1

Weight gain of male (A) and female (B) C56Bl/6 mice fed a standard chow control (LFD), and (HFD) and kefir fractions on a HFD background (CFK, HK, and PK1). Data are expressed as mean values with their standard errors (n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.

Figure 2

Concentration of total plasma cholesterol (A,D), HDL cholesterol (B,E), and non-HDL cholesterol (C,F) in male/female C57Bl/6 mice fed a control diet or high-fat diet supplemented with milk or different kefir preparations for 8 weeks. Data are expressed as mean values with their standard errors (n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.

Figure 3

Concentration of liver cholesterol (A,C) and triglycerides (B,D) in male/female C57Bl/6 mice fed a control diet or high-fat diet supplemented with milk or different kefir preparations for 8 weeks. Data are expressed as mean values with their standard errors (n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.

Figure 4

Concentration of plasma VCAM-1 (A,B) and CRP (C,D) in male/female C57Bl/6 mice fed a control diet or high-fat diet supplemented with milk or different kefir preparations for 8 weeks. Data are expressed as mean values with their standard errors (n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.

Figure 5

Alpha diversity metrics of the cecal microbiota in male/female C57Bl/6 mice fed a control diet or high-fat diet supplemented with milk or different kefir preparations for 8 weeks. (A) Chao1 and PD indices indicated higher microbial diversity and richness in the LFD group (p < 0.05) and lower diversity in the Kefir group (p < 0.05) compared to HFD, CFK and HK groups for females with no difference with Shannon diversity index other than LFD maintained higher diversity. (B) No difference was found in male mice after kefir consumption for Chao1, Shannon or PD metrics compared to HFD group (n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.

Figure 6

Principle coordinate analysis using unweighted and weighted UniFrac distance metric of the cecal microbiota of male/female C57Bl/6 mice fed a control diet or high-fat diet supplemented with milk or different kefir preparations for 8 weeks. A major shift between LFD and HFD groups was determined in both (A) female and (B) male mice with no difference between HFD and kefir consumption groups (CFK, HK and Kefir; n = 7–8). Means that do not share a letter are significantly different (p < 0.05). LFD, mice fed a low fat diet supplemented with freeze-dried milk for 8 weeks; HFD, mice fed a high fat diet supplemented with freeze-dried milk for 8 weeks; PK1, mice fed a high fat diet supplemented with freeze-dried pitched kefir for 8 weeks; CFK, mice fed a high-fat diet supplemented with freeze-dried cell-free kefir; HK, mice fed a high-fat diet supplemented freeze-dried heat-treated kefir.