*3.4. Animals and Experimental Design*

Adult male Sprague-Dawley rats (220–240 g) were provided by Experimental Animal Center of Shandong University. The animals were maintained on a 12 h dark/light cycle at about 22 ◦C and relative humidity (60–70%), allowed free access to standard rat chows and water during the experiments. The experiments were performed in complete compliance with the National Guide for the Care and Use of Laboratory Animals, and were approved by the Experimental Animal Ethics Committee of Institute of Oceanology, Chinese Academy of Sciences, China (approval code SCXK(Lu)20190002).

The adriamycin-induced nephrotic syndrome model in rats was performed according to Bertani et al. protocol [2]. A single dose of adriamycin (6.0 mg/kg body weight) was injected via the femoral vein to induce nephrotic syndrome (NS) model. After injection of adriamycin, 24 h urinary protein content was measured once a week. Two weeks following the adriamycin injection, proteinuria was detected, then the rats were randomly divided into six groups (*n* = 10): Group 1 was the model group which consisted of NS rats; Group 2 consisted of 10 NS rats and the rats were treated with daily oral gavage of fucoidan at dosage of 100 mg/kg body weight; Group 3, 4, and 5 consisted of NS rats and were treated daily oral gavage of LMWF at dosage of 100, 50, and 25 mg/kg body weight, respectively. Group 6 was the positive control group, and NS rats in this group were administrated with dexamethasone acetate at dose of 0.1 mg/kg body weight. Group 7 was the normal group (*n* = 10) and rats were orally administrated with 10 mL/kg/d of saline. All rats were raised in the same environment and were allowed unlimited access to water and conventional rat chow during the experiments. During the experiments, the animals were weighed twice a week, the general state of animals was recorded every day, including body hair, stool, and mental state.

To measure urinary protein levels, the rats were placed in individual metabolic cages for 24 h urine collection once a week.

On the thirtieth experimental day the animals were anaesthetized by ether and blood samples were taken from the eye-pit of rats. After the blood sample was taken, the rats were killed and the kidneys were weighted and taken for routine histological examination.
